Abstract Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype characterized by the absence of targetable receptors and limited therapeutic options. Its inherently low immunogenicity necessitates strategies to enhance its susceptibility to immune attack. This study aimed to enhance the antitumor immune response by inducing downregulation of the programmed death ligand 1 (PD-L1), thereby increasing cancer cell recognition and activating tumor-specific immunity. To this end, a targeted solid lipid nanoparticle (tSLN) system was developed for the precise delivery of CRISPR/Cas9 plasmids to achieve functional PD-L1 downregulation. The SLN formulation comprised a cetyl palmitate core stabilized with cationic and PEGylated lipids. Functionalization with the tumor-penetrating peptide iRGD yielded tSLN, which demonstrated good colloidal stability, serum protection of the CRISPR cargo, and minimal cytotoxicity. In vitro studies in two breast cancer subtypes, the luminal MCF-7 and the triple-negative 4T1 line, confirmed greater transfection efficiency and PD-L1 knockdown by tSLN:Crispr/PD-L1 relative to non-targeted controls. In an orthotopic 4T1 mouse model, systemic administration of tSLN:Crispr/PD-L1 significantly inhibited tumor growth and metastasis while enhancing T cell infiltration, and was more effective than standard chemotherapy (paclitaxel) in this model. The targeted approach also exhibited a favorable short-term safety profile, with no organ toxicity detected over the treatment period. These findings underscore the potential of SLN-mediated CRISPR delivery as an immunomodulatory strategy for breast cancer, and particularly for TNBC.
Abstract Quorum sensing (QS) is a bacterial communication mechanism that regulates collective behaviors such as virulence, motility, and biofilm formation. Consequently, materials capable of interacting with molecules associated with QS signaling pathways are of increasing interest as potential tools for studying and modulating bacterial communication processes. In this study, molecularly imprinted polymers (MIPs) were designed and synthesized using (R/S)-2,2-dimethyl-1,3-dioxolane-4-methanamine, an autoinducer-2 (AI-2)-inspired surrogate template to investigate molecular recognition of structures relevant to bacterial signaling systems. Five bulk MIPs were prepared by thermal radical polymerization using methacrylic acid, 4-vinylbenzoic acid, itaconic acid, trifluoromethacrylic acid, and acrylic acid as functional monomers. The resulting polymers were evaluated in terms of adsorption capacity, imprinting factor, and selectivity toward the template and structurally related compounds using liquid chromatography coupled with mass spectrometry (LC−MS)-based binding studies. Among the synthesized materials, the polymer prepared from methacrylic acid exhibited the highest specificity, with an imprinting factor of 3.10 at 50 μg L−1 and up to 19.14 at 2 μg L−1 of template concentrations, indicating strong concentration-dependent recognition. Computational modeling using density functional theory, molecular mechanics, and molecular dynamics simulations was employed to elucidate the molecular interactions governing template recognition and cavity formation. Theoretical analyses revealed that multiple hydrogen bonding, hydrophobic interactions, and electrostatic contacts contribute to stable analyte-polymer binding, supporting the experimentally observed selectivity. Selectivity experiments demonstrated preferential adsorption of the template and its closest structural analogue relative to several structurally related amine-containing compounds. Biological evaluation using the Chromobacterium violaceum model revealed that both molecularly imprinted and non-imprinted polymers reduced violacein production to varying degrees, indicating modulation of a QS-regulated phenotype under the experimental conditions employed. Collectively, these findings demonstrate the feasibility of using molecular imprinting strategies to generate polymeric materials capable of recognizing an AI-2-inspired surrogate template and influencing a QS-regulated biological response, highlighting their potential for development as innovative materials for antivirulence strategies against bacterial infections.
Abstract Nanoparticle barcoding is a powerful approach to accelerate preclinical screening of nanotherapeutics by enabling rapid collection of high-volume data using fewer biological resources, with reduced experimental labor and fewer confounding variables across experiments. We previously introduced aryl halide-based tags (halocodes) as a barcoding technology with sensitive gas chromatography−mass spectrometry (GC−MS) detection, enabling nanoparticle tracking with minimal halocode incorporation and without measurably altering nanoparticle physicochemical properties. Building on this foundation, in this work, we developed a next generation of alkoxy-modified halocodes with tunable chemical properties that enhance encapsulation and stability within nanomaterials and extend the halocoding platform to lipid-based systems. By systematically varying halocode chemical structure and nanoparticle synthesis parameters, we established design rules governing halocode loading and retention in both polymeric nanoparticles and liposomes. We then leveraged next generation halocodes to perform a pooled in vitro screen of a mixed-core PLGA and liposomal-based nanoparticle library to identify nanoparticle core and surface chemistries with the highest affinity for ovarian cancer cells. Together, next generation halocodes provide a more modular and broadly compatible barcoding toolkit that simplifies pooled screening and supports quantitative evaluation of diverse nanocarrier libraries across biological settings.
Abstract The solubility advantage of an amorphous drug can be realized only when it releases at a sufficiently rapid rate from its amorphous solid dispersion (ASD). Herein, we evaluated the impact of a plasticizer, glyceryl tributyrate (GTB), on the release of a poorly soluble drug with a high glass transition temperature (Tg). Cyclosporine has a Tg of 124 °C and was formulated as an ASD with hydroxypropyl methylcellulose acetate succinate (HPMCAS). Binary and ternary ASDs were prepared using solvent evaporation, their Tgs were measured using differential scanning calorimetry, and surface area normalized release rates were determined. The drug-rich phase formed above the amorphous solubility was characterized for composition and size. Drug flux following release from the ASD was measured across Caco-2 cell monolayers. Cyclosporine released at a substantially faster rate from ternary ASDs containing GTB as compared to the corresponding binary ASD. This improvement was attributed to a decrease in Tg upon plasticization due to GTB increasing molecular mobility. Addition of GTB, however, reduced the amorphous solubility of cyclosporine resulting in a decrease in the transport rate across the Caco-2 cell monolayer. These observations may have implications for drug absorption. Ultimately, it is expected that the in vivo performance of these ternary ASDs would be determined by the interplay between the enhanced release rate and the decreased permeation rate of the drug.
Abstract In high-drug-loaded amorphous solid dispersions, effective stabilization requires polymers that can hinder nucleation as well as crystal growth. We hypothesized that combining two polymers with complementary stabilization mechanisms would enhance physical stability more effectively than the individual polymers. Evaluation of single-polymer systems at a polymer loading of 20% w/w showed that HPMCAS (hydroxypropyl methylcellulose acetate succinate) had the highest thermodynamic miscibility with carbamazepine (CBZ), whereas PVP (polyvinylpyrrolidone) and PVPVA (PVP/vinyl acetate) dispersions had the longest α-relaxation time. Based on these findings, ternary systems combining 80% w/w CBZ, 10% w/w HPMCAS, and 10% w/w of either PVP or PVPVA were evaluated. The ternary systems exhibited longer crystallization onset times than the corresponding single-polymer systems at the same total polymer loading. The improved stability was due to a synergistic mechanism where HPMCAS provided resistance to nucleation by increasing the activation energy barrier, while PVP/PVPVA kinetically lowered the attempt jump frequency to suppress both nucleation and crystal growth. This dual-polymer approach provided a basis for designing amorphous solid dispersions with high drug loading and improved physical stability.
Abstract Gastric cancer is a highly prevalent malignant tumor of the digestive system worldwide. Although andrographolide (AG) and coixol (CX) possess potential anti-gastric cancer effects, their clinical application is restricted by low bioavailability and poor targeting ability. Based on the meridian tropism theory of traditional Chinese medicine and the concept of “integrated drug and carrier”, this study constructed Bletilla striata polysaccharide-vitamin E succinate (BSP-VES) polymeric micelles co-loaded with AG and CX (AG&CX@BSP-VES) for synergistic targeted therapy of gastric cancer. In this study, AG&CX@BSP-VES was first systematically characterized in vitro to clarify its physicochemical properties, drug release behavior and hemolysis profile. Subsequently, using subcutaneous and orthotopic gastric cancer mouse models as research objects, in vitro cellular uptake and in vivo tumor targeting ability efficacy and biological safety of this drug delivery system were further evaluated. Results showed that the micelles were regularly spherical with a particle size of approximately 105 nm, exhibited favorable biological safety and asynchronous differential release characteristics, and could be specifically internalized by cells in vitro (which could be blocked by free BSP). In vivo, the accumulation of micelles in tumor masses was significantly higher than that in normal gastric tissues. Moreover, the cytotoxicity of the micelles against SGC-7901 tumor cells was remarkably higher than that of free AG or CX alone, which was attributed to enhanced cellular drug uptake and the synergistic effect between AG and CX. This study provides novel insights for the precise delivery of active ingredients from traditional Chinese medicine and individualized therapy of gastric cancer.
Abstract Intracellular infections impose fundamental physicochemical barriers that can disrupt antibiotic synergy observed under well-mixed in vitro conditions. Mycobacterium abscessus, a macrophage-resident pathogen, exemplifies this challenge, as co-administered drugs must traverse multiple membrane barriers and achieve coordinated intracellular exposure. The combination of clofazimine (CFZ) and amikacin (AMK) exhibits robust, time-dependent synergy in broth, yet their divergent physicochemical properties (lipophilic versus hydrophilic) predict differential intracellular access. Here, we evaluated whether formulation architecture can mitigate this spatial and temporal separation. CFZ and AMK were engineered either as physical mixtures of micronized powders or as spray-dried composite particles across multiple drug ratios and compared in terms of aerosol performance and intracellular efficacy in a THP-1 macrophage infection model. Neither drug alone achieved meaningful intracellular killing despite activity in broth. In contrast, combination efficacy was strongly formulation-dependent and exhibited a non-monotonic relationship with composition. Physical mixtures were most effective at CFZ-rich ratios, whereas spray-dried composites achieved superior killing at an intermediate mass ratio (1:4 CFZ/AMK) and displayed a delayed-onset kinetic profile consistent with architecture-dependent temporal drug availability. Aerosol studies showed that composite particles generally improved dispersibility and reduced bulk aerodynamic fractionation between the two drugs relative to physical mixtures, although this effect was condition-dependent. Together, these findings indicate that intracellular efficacy is governed not solely by drug identity or dose, but by formulation-controlled spatial and temporal drug presentation. This work establishes formulation architecture as a critical design variable for translating antibiotic synergy into biologically constrained environments.
Abstract Gemcitabine (GEM) is a first-line therapeutic option for pancreatic cancer; however, it has low efficacy due to rapidly developed drug resistance and severe dose-limiting myelosuppression. To enhance its therapeutic effect, this study developed a novel peptide−drug conjugate using a CXCR4 antagonistic peptide as the targeting head and gemcitabine as the drug payload, based on the characteristics of pancreatic tumor cells highly expressing CXCR4 that mediates immunosuppression and tumor progression through interaction with its specific ligand CXCL12. The therapeutic effect of the conjugate (P12-GEM) was investigated using pancreatic ductal adenocarcinoma cell lines and an orthotopic pancreatic cancer mouse model. Its myelosuppressive effect was assessed from the perspective of hematological toxicity profiles. The results showed that P12-GEM maintained a cell-killing capability comparable to that of GEM while effectively inhibiting the phosphorylation of Erk and P38, thereby reducing CXCL12-mediated tumor cell migration and adhesion to stromal cells. In a tumor-bearing mouse model, P12-GEM demonstrated superior antitumor efficacy compared to GEM and significantly extended animal survival. Moreover, P12-GEM reduced the proportion of tumor-associated macrophages and increased the infiltration of CD8+ T cells in the tumor microenvironment without reducing platelet and white blood cell counts in the peripheral blood. In summary, P12-GEM possesses dual functions of CXCR4 antagonism and tumor cell killing, contributing to reversal of the immunosuppressive microenvironment and alleviation of myelotoxicity.
Abstract 18β-Glycyrrhetinic acid (GA) exhibits excellent anti-inflammatory and antioxidant activities, and has attracted considerable attention in the field of cerebral ischemia−reperfusion (I/R) injury therapy. However, it is plagued by problems of low bioavailability and dose-dependent toxicity. As a drug delivery platform for the treatment of ischemic stroke, microenvironment-responsive liposomes can cross the impaired blood−brain barrier, efficiently deliver drugs to the lesion sites, and improve therapeutic efficacy. Herein, to address the challenges posed by the highly oxidative-stress microenvironment of I/R and the intrinsic physicochemical properties of GA, we encapsulated GA into liposomes and incorporated thioketal (TK) linkages into the phospholipid bilayer to fabricate GA@TK-Lip. GA@TK-Lip can effectively target damaged neurons, attenuate neuroinflammation, scavenge excessive ROS, ameliorate cellular oxidative stress via the Nrf2/HO-1 pathway, reduce neuronal apoptosis and cerebral infarct volume to 43.2% and 23.3% of that in the MCAO group, and restore neurological function. Collectively, this study provides novel insights into the development of promising therapeutic strategies for ischemic stroke.
Abstract Numerous drug candidates showed great promise in experimental and preclinical investigations but failed in clinical trials. One cause to this problem may be nonspecific drug biodistribution, leading to adverse effects in healthy organs. Drug delivery technologies are developed to overcome this exact problem by navigating drugs more selectively to the disease sites. Therefore, they hold the promise to revive at least some failed drugs. However, it is challenging to identify such candidates revivable by drug delivery: first, there lacks systemic survey and documentation of drug biodistribution in patients during the trials; second, clinical trial results, especially the failed ones, are often reported in unstructured and/or inconsistent language. Here, we harnessed the recent advances in artificial intelligence (AI) agents and large language models (LLMs) to tackle with the second problem. An OpenAI-based framework was developed to systemically analyze terminated trials on ClinicalTrials.gov. This framework integrated structured extraction of clinical trial information, disease-site inference with evaluation based on predefined screening criteria, which is unwanted biodistribution (UB failure) reflected by the difference in the anatomical locations between adverse effects and targeted diseases. We also paired this with the analysis of drug types and physicochemical properties (e.g. molecular weight and hydrophobicity). Using a validation set, the model achieved accuracies of 89.69% for UB failure classification, 91.75% for small molecule drug classification, and 88.66% for hydrophobicity classification. We then applied this analysis to all terminated ClinicalTrials.gov trials, which identified 817 trials possibly failed due to UB, and top ten small molecule drugs likely suitable for our peptide-guided delivery platform. Overall, this study may provide a new direction to apply AI technologies for drug development, and inspire future investigations of clinical reports and public information from the perspective of delivery technology applications.
Abstract Fibroblast activation protein (FAP)-targeted radioligand therapy is often limited by insufficient tumor uptake and retention. We designed and synthesized two novel aryl fluorosulfate-based covalent FAP-targeted radioligands, [177Lu]Lu-DOTA-mFS-FAP-2286 and [177Lu]Lu-DOTA-mFS-KERERG-FAP-2286, to enhance tumor retention and therapeutic efficacy. Two FAP-targeted radioligands were synthesized, radiolabeled with 177Lu, and evaluated for radiochemical stability. Cellular uptake and internalization were assessed in HEK293-huFAP cells. Biodistribution and serial SPECT imaging were performed in tumor-bearing mice to evaluate tumor targeting and retention. Therapeutic efficacy was investigated in murine xenograft models, and the toxicity of [177Lu]Lu-DOTA-mFS-KERERG-FAP-2286 was assessed. A pilot clinical study including three patients with solid tumors was conducted to preliminarily evaluate the therapeutic potential. All radioligands were successfully labeled with radiochemical purity >95% and showed favorable stability. Cellular assays confirmed the high target specificity. In tumor-bearing mice, [177Lu]Lu-DOTA-mFS-KERERG-FAP-2286 demonstrated significantly higher tumor uptake at 1 h after injection than [177Lu]Lu-DOTA-FAP-2286 and [177Lu]Lu-DOTA-mFS-FAP-2286 (p < 0.01). Treatment with [177Lu]Lu-DOTA-mFS-KERERG-FAP-2286 significantly inhibited tumor growth and showed superior therapeutic efficacy compared with [177Lu]Lu-DOTA-FAP-2286 (p < 0.05). No treatment-related adverse events were observed in the three patients. Primary tumors and metastatic lesions demonstrated high tracer uptake with prolonged retention up to day 10 after injection. [177Lu]Lu-DOTA-mFS-KERERG-FAP-2286 showed favorable tumor targeting, prolonged retention, and potent therapeutic efficacy, supporting its further clinical development for FAP-targeted radioligand therapy (RLT).
Abstract Reactive oxygen species (ROS) are significantly elevated in tumor cells, providing an intrinsic trigger for selective drug activation and tumor-specific therapy. However, commonly used ROS-responsive linkers, such as thioketals and boronates, often suffer from limited sensitivity or poor physiological stability, and the development of new ROS-responsive motifs remains scarce. Herein, we develop a hydrazide-derived diazaborine (DAB) scaffold and establish it as a general ROS-responsive linker for small-molecule prodrugs, combining physiological stability with efficient ROS responsiveness. Based on this platform, we construct a DAB-derived ROS-responsive copper complex small-molecule prodrug, DDABD@Cu, for tumor-specific therapy. DDABD@Cu integrates a DAB-caged diethyldithiocarbamate (DTC) moiety with an inert copper donor, in which the DAB group effectively blocks the copper-chelating activity of DTC, enabling inert co-delivery of both components. Upon ROS activation, DTC is released to initiate copper transfer, generating cytotoxic Cu(DTC)2 in situ to induce proteotoxic stress via ubiquitinated protein accumulation while concomitantly producing quinone methide (QM) to deplete glutathione (GSH) and amplify oxidative stress. This cascade improves antitumor efficacy against non-small cell lung cancer while limiting off-target toxicity. Overall, this work provides a feasible strategy for designing ROS-responsive and copper complex prodrugs.
Rapid crystallization of an active pharmaceutical ingredient can suppress its observable supersaturation and negate this potential advantage of the amorphous form. Excipients added to formulations can prolong supersaturation, but reduce drug loading and obscure the intrinsic dissolution behavior of the drug in testing. Here, we describe spring-and-parachute dissolution behavior in fasted-state simulated intestinal fluid of an archetypal poorly soluble and strongly crystallizing drug, griseofulvin (GSF), in the form of excipient-free, pure, amorphous nanopowders generated using a novel, single-step, solvent-free technique of organic vapor jet desublimation. Using experiments and modeling, we establish how initial dissolution rate, peak concentration, and area under the concentration-time curve scale with dose, distinguishing between dose regimes in which supersaturation behavior is monotonic and nonmonotonic. The roles of local surface crystallization, particle aggregation, and nonequilibrium crystallization at high supersaturation are elucidated, providing a framework for attaining excipient-free supersaturation and guiding the development of improved dissolution models.
The high expression of transferrin receptors on the surface of cancer cells makes them a potential target for drug internalization, which has become a golden opportunity in targeted cancer therapy. Carvacrol, a naturally occurring phenolic compound present in essential oils, has attracted considerable attention for its pronounced anticancer activity against diverse human malignancies. Nevertheless, its therapeutic application in vivo is constrained by intrinsic physicochemical and pharmacokinetic limitations, including poor water solubility, suboptimal bioavailability, and rapid systemic elimination. To address these challenges, the present study highlights the fabrication of transferrin-targeted TPGS-conjugated carvacrol (CVC)-loaded polydopamine nanoparticles (CVC-PDA-TPGS-Tf NPs), with exploration of the antitumor efficacy of nanocarrier-mediated carvacrol, deploying a murine triple-negative breast cancer (TNBC) tumor xenograft model and a human TNBC three-dimensional (3D) spheroid model, as 3D cell culture technologies more thoroughly bear a resemblance to in vivo cell environments to accommodate improved precision in drug discovery. This work demonstrates the successful creation of nanoconjugate CVC-PDA-TPGS-Tf NPs (∼112 nm in size) and their thorough physicochemical characterization after synthesis. Intraperitoneal administration of CVC-PDA-TPGS-Tf NPs substantially inhibited tumor growth in the 4T1 xenograft model. Carvacrol effectively disintegrates MDA-MB-231 spheroids, illustrating a proportionately higher uptake of carvacrol nanoconjugates by these spheroids. The molecular docking studies also pinpointed the interaction of potential cancer biotargets with carvacrol in estrogen- and progesterone-receptor-positive breast cancer cells, in addition to its mentioned therapeutic efficiency against TNBC. In conclusion, these findings support CVC-PDA-TPGS-Tf as a promising platform for TNBC and potentially hormone receptor-positive breast cancers, warranting further mechanistic and translational studies.
The use of natural killer (NK) cells for the treatment of solid tumors such as cholangiocarcinoma has been hindered by challenges related to cell production, persistence and trafficking, as well as by reduced efficacy within an immunosuppressive tumor microenvironment. Antitumor immunity can be reduced by the interaction of NK cell expressed programmed cell death protein 1 (PD1) with ligands on tumor and other cells within the tumor microenvironment. The antitumor effect of NK cells is enhanced in cells engineered to express a truncated PD1 without an intracellular domain. The cytolytic potential and therapeutic efficacy of extracellular vesicles (EVs) derived from human NK cells expressing truncated PD1 was evaluated as a cell-free therapeutic to overcome some limitations faced by cell therapies. We demonstrate the feasibility of efficient expansion, production and isolation of therapeutic vesicles from human NK cell lines and from engineered NK92 cells expressing truncated PD1. The cytotoxic efficacy of these therapeutic vesicles was validated in both monolayer tumor cell cultures and in multicellular tumor spheroids. Concomitant administration of gemcitabine upregulated NKG2D ligands on tumor cells and increased susceptibility to NK cell and to NK cell derived EV mediated killing. These findings position engineered NK cell derived-EVs as a scalable and promising cell-free immunotherapeutic for cholangiocarcinoma.
Human serum albumin (HSA) plays a key role in regulating the transport and bioavailability of therapeutic molecules, making protein-ligand interaction studies essential for assessing biomedical compatibility. In this work, the interaction of cotarnine and its derivatives with HSA was systematically investigated using spectroscopic, thermodynamic, and molecular docking approaches, along with evaluation of antibacterial, wound healing, and hemostatic activities. Spectroscopic analyses confirmed efficient binding of the derivatives to HSA without disrupting its native secondary structure, while thermodynamic studies revealed spontaneous, enthalpy-driven interactions dominated by noncovalent forces. Docking studies identified favorable binding orientations within HSA binding pockets, complementing the experimental results. All derivatives of cotarnine show greater binding and biological activity, such as wound healing and hemostatic, than cotarnine. Among the derivatives, L2 exhibited the strongest binding affinity and superior biological performance, which can be attributed to enhanced hydrophobic interactions arising from its chloro-substituted aromatic moiety. Overall, the combined biophysical, computational, and biological findings highlight cotarnine derivatives, particularly L2, as promising multifunctional candidates for wound-healing and related biomedical applications.
Organic anion and cation transporters (OATs and OCTs), members of the solute carriers (SLC)22 family, are critically involved in the transport of endogenous metabolites and xenobiotics across key barriers in the kidney, liver, and choroid plexus. While their role in small-molecule drug pharmacokinetics, drug-drug interactions, and clinical issues is well established, their potential interaction with nanomaterials remains a largely unexplored frontier. This is particularly significant given that the long-term retention of nanomaterials in vivo poses a major challenge to their clinical translation. We hypothesize that OATs and OCTs recognize specific nanomaterials as substrates, thereby mediating a novel and active elimination pathway that transcends conventional size-dependent filtration. This review systematically delineates the structural and functional basis of major OAT/OCT subtypes (e.g., OAT1-3, URAT1, OCT1-3) to establish a conceptual framework and then integrates this with current clinical applications and a brief discussion on the primary biological elimination routes of nanomaterials. By bridging these fields, this review aims to prompt a paradigm shift in nanomedicine design, advocating for the rational engineering of nanomaterial properties (e.g., size, charge, surface, shape) to engage with these transport systems. Ultimately, leveraging OAT/OCT pathways may unlock novel strategies to precisely control nanomedicine distribution and enhance clearance, thereby accelerating safe and effective clinical translation.
Macropa has formed more stable complexes with 225Ac, although DOTA has been extensively used as a chelator for 225Ac. This study aimed to design and synthesize two novel 225Ac-labeled RGD peptides, [225Ac]Ac-Macropa-c(RGDyK) ([225Ac]1) and [225Ac]Ac-Macropa-[c(RGDyK)]2 ([225Ac]2), and to evaluate their potential for targeted alpha therapy (TAT) of glioblastoma. Further, whether combination therapy with homoarginine (hArg), a modulator of lysosomal function, could further improve therapeutic efficacy was investigated. In Colon-26 cells, [225Ac]2 demonstrated significantly higher cellular uptake and cytotoxicity than [225Ac]1. In Colon-26 tumor-bearing mice, [225Ac]2 exhibited higher tumor accumulation and significant tumor growth inhibition. In GL261 glioblastoma cells, [225Ac]2 showed comparable cellular uptake to that in Colon-26 cells, and combination with hArg in vitro improved its cytotoxicity. However, no significant difference in therapeutic efficacy was observed between the hArg-treated and control groups in the orthotopic GL261 glioblastoma mouse model. The results of this study indicate the potential of 225Ac-labeled RGD peptides incorporating Macropa as a chelator as promising agents for TAT. However, the further optimization of combination strategies and pharmacokinetics is required.
Porphyrin bimetallic metal-organic frameworks (BMOFs) have promising applications in photodynamic therapy (PDT) and chemodynamic therapy (CDT) for tumor therapy. However, their therapeutic effect is restricted by the insufficient reactive oxygen species generated by BMOFs. Herein, a nanobomb, Fe-TCPP(Mn)/DOX/PEG/Apt-M (FTMDPA), with pH-responsive properties, was designed for fluorescence imaging-guided triple-action tumor annihilation. First, a novel Fe/Mn porphyrin BMOF, Fe-TCPP(Mn) (FTM), was assembled by a solvothermal method to achieve enhanced tumor therapy through ROS amplification. Under 660 nm laser irradiation, TCPP(Mn) could act as a photosensitizer to activate PDT. Significantly, TCPP(Mn) and Fe3+ exhibit peroxidase-like and Fenton-like activities, catalyzing the generation of •OH from H2O2, which effectively amplifies CDT. Besides, FTMDPA exhibits pH-responsive DOX release capability, which could achieve chemotherapy (CHT), and could efficiently accumulate at the tumor site with the aid of an aptamer. Importantly, FTMDPA exhibits fluorescence imaging, which has potential applications in detection and image-guided precision therapy. Therefore, the designed nanobomb integrates targeted delivery, pH responsiveness, fluorescence imaging, and amplified PDT-CDT-CHT triple-action tumor annihilation into a single system, providing a promising strategy for tumor diagnosis and treatment.
Necroptosis is a regulated, pro-inflammatory form of programmed cell death implicated in degenerative and inflammatory diseases, with RIPK1 acting as a key upstream regulator. However, the utility of existing RIPK1 PET radiotracers, as well as various RIPK1 inhibitors, remains uncertain, as it is unclear whether they reflect total protein abundance or phosphorylation-dependent activation (pRIPK1), limiting accurate assessment of in vivo signaling. Unlike previously reported RIPK1 PET tracers that primarily reflect total RIPK1 distribution, we investigated whether [18F]RIPA56-3-F ([18F]1) uptake is associated with RIPK1 activation-related signaling in vivo. [18F]1 was synthesized via copper-mediated radiofluorination, and its radiochemical properties were characterized. In vitro uptake studies were performed in ARPE-19 cells under sodium iodate-induced stress to assess the relationship between [18F]1 uptake, pRIPK1 activation, and total RIPK1 expression. In vivo dynamic PET/CT imaging was conducted to evaluate pharmacokinetics and biodistribution. A retinal injury model induced by sodium iodate was used for time-course imaging and quantitative analysis of radiotracer uptake. Pharmacologic validation was performed using the RIPK1 inhibitor necrostatin-1s. Ex vivo analyses, including immunohistochemistry, were conducted to correlate PET findings with pRIPK1 expression. [18F]1 was obtained with a non-decay-corrected radiochemical yield of 37.7 ± 6.1% (n = 27), radiochemical purity >99%, and stability up to 4.5 h. In ARPE-19 cells, [18F]1 uptake increased in parallel with early pRIPK1 activation and decreased at later time points despite sustained total RIPK1 expression. In vivo dynamic PET demonstrated favorable pharmacokinetics, with a pseudo-steady state observed at 60-80 min postinjection. Time-course analysis revealed that tracer uptake peaked at early stages (7-12 h) following sodium iodate administration and declined thereafter, consistent with transient pRIPK1 activation. In the retinal injury model, ocular uptake of [18F]1 was significantly higher than in controls (3.38 ± 0.45 vs 1.52 ± 0.21%ID/g; P < 0.0001) and was significantly reduced by necrostatin-1s treatment (P = 0.0096). PET signal was detectable prior to overt structural degeneration and correlated with increased pRIPK1 expression. [18F]1 uptake was more closely associated with pRIPK1-related signaling activity than with total RIPK1 expression, enabling noninvasive imaging of necroptosis-associated signaling in vivo. This activation-state imaging approach allows detection of early molecular events preceding structural damage and supports the use of [18F]1 as a potential pharmacodynamic biomarker for assessing RIPK1 pathway engagement and therapeutic modulation.
