Molecular Pharmaceutics[JOURNAL]
Sun
200 papers
The Use of Glycine-Containing Peptide-Based Chelators for Labeling with Tc Improves the Imaging Properties of EpCAM-Targeting Designed Ankyrin Repeat Ec1.
Deyev S, Fominykh A, Varvashenya R … +13 more , Yanovich G, Bodenko V, Plotnikov E, Tretyakova M, Eskova D, Zhelchan R, Schulga A, Konovalova E, Ziganshin R, Orlova A, Belousov M, Tolmachev V, Larkina M
Mol Pharm · 2026 Apr · PMID 41780028 · Publisher ↗
Noninvasive radionuclide imaging of epithelial cell adhesion molecule (EpCAM) expression in lung, ovarian, breast, kidney, and other cancers can stratify patients for EpCAM-targeted therapy. The constructed scaffold prot...
Noninvasive radionuclide imaging of epithelial cell adhesion molecule (EpCAM) expression in lung, ovarian, breast, kidney, and other cancers can stratify patients for EpCAM-targeted therapy. The constructed scaffold proteins, designed ankyrin repeat proteins (DARPins), are highly specific high-affinity probes for radionuclide imaging. A clinical study demonstrated that the anti-EpCAM DARPin [Tc]Tc-(HE)-Ec1 showed precise EpCAM imaging at 2, 4, and 6 h after injection in patients with nonsmall cell lung cancer. However, a noticeable accumulation in healthy organs has prompted the development of new Ec1-based agents with improved biodistribution properties. In addition, it would be desirable to substitute a labor-intensive labeling procedure. The purpose of this study was to test the hypothesis that the use of Gly-Gly-Gly-Cys (GC) or Glu-Glu-Glu-Cys (EC) peptide chelators placed at the C-terminus of DARPin for labeling with Tc (V) could improve the image contrast and biodistribution of Ec1. The radiochemical yield of the new variants exceeded 95%. The labeled proteins specifically bound to human EpCAM-expressing cancer cell lines with affinities of 8-10 nM. The biodistribution of [Tc]Tc-Ec1-GC and [Tc]Tc-Ec1-EC in mice was compared with the biodistribution of clinically tested [Tc]Tc-(HE)-Ec1 in a Nu/j mouse model with SKOV-3 xenografts. The new variants specifically accumulate in human xenografts with EpCAM expression. The accumulation of new variants in healthy organs (liver, salivary glands, spleen, and stomach) was reduced compared to [Tc]Tc-(HE)-Ec1. [Tc]Tc-Ec1-GC provided the best imaging contrast and is suitable for clinical testing.
Nebulized Inhalation of ROS-Responsive Dexamethasone-Loaded Liposomes Enhances Therapeutic Efficacy against Pulmonary Fibrosis.
Sun J, Jiao Y, Jin Y … +4 more , Zhao Z, Xing Y, Yan Y, Quan J
Mol Pharm · 2026 Apr · PMID 41778757 · Publisher ↗
This study aimed to develop and evaluate reactive oxygen species (ROS)-responsive (Thioketal-grafted DSPE-PEG2000, DTP) liposomes loaded with dexamethasone (DTP@DEX-LP) for aerosol inhalation to achieve targeted drug del...
This study aimed to develop and evaluate reactive oxygen species (ROS)-responsive (Thioketal-grafted DSPE-PEG2000, DTP) liposomes loaded with dexamethasone (DTP@DEX-LP) for aerosol inhalation to achieve targeted drug delivery and controlled release in the pulmonary fibrosis microenvironment. DTP@DEX-LP was prepared and optimized. The liposomes were characterized for particle size, zeta potential, encapsulation efficiency (EE), and morphology. Their aerosol performance, ROS-responsive drug release, and cellular uptake were assessed . pulmonary deposition, pharmacokinetics, and antifibrotic efficacy were evaluated in a bleomycin-induced mouse model, alongside safety profiling. The optimized DTP@DEX-LP exhibited a uniform particle size of ∼ 115 nm, a high EE of >82%, and desirable aerosol properties (FPF ∼ 50%, MMAD ∼ 4.9 μm). The formulation demonstrated ROS-triggered drug release and enhanced cellular uptake . Following inhalation, DTP@DEX-LP significantly prolonged lung retention and reduced systemic exposure of DEX compared to controls. In the fibrosis model, DTP@DEX-LP treatment yielded superior therapeutic outcomes, markedly improving survival, and reducing collagen deposition. It also showed a notably improved safety profile, with reduced hepatotoxicity compared to intravenous DEX. The ROS-responsive liposomal system represents a promising inhaled platform for the precise treatment of pulmonary fibrosis, effectively enhancing the therapeutic index of dexamethasone by simultaneously improving its efficacy and mitigating systemic toxicity.
Investigation of Radiolabeled KISS1R Ligands as Promising Tools for Diagnosis and Treatment of Triple-Negative Breast Cancer.
Taş H, Schäfer M, Shuja-Uddin A … +13 more , Bauder-Wüst U, Kovacs Dos Santos L, Bartnitzky L, Oden F, Platzk M, König T, Rüther PL, Pook E, Dvořáková Bendová K, Nový Z, Petřík M, Hagemann UB, Benešová-Schäfer M
Mol Pharm · 2026 Apr · PMID 41774810 · Full text
Kisspeptins (KPs) and their receptor (KISS1R) promote metastasis and tumor progression in various cancers such as triple-negative breast cancer (TNBC). Targeting KISS1R holds great promise for molecular imaging and targe...
Kisspeptins (KPs) and their receptor (KISS1R) promote metastasis and tumor progression in various cancers such as triple-negative breast cancer (TNBC). Targeting KISS1R holds great promise for molecular imaging and targeted radionuclide therapy of aggressively disseminated cancers. First ligand-based approaches using Ga-68/Lu-177-labeled KPs (KP-10, KP-54) have demonstrated feasibility but suffer from proteolytic degradation and low uptake in KISS1R positive tumors. However, lead structure optimization alone is insufficient, as KISS1R biology remains unexplored in a radiotheranostic context. In this study, -terminally functionalized conjugates of KP-10, KP-54, and the hybrid peptide KiSS-34 (AMBA-2-Nal-Gly-Leu-Arg-Trp-NH), including scrambled controls, were synthesized in high purity (≥95%) for comparative studies. The conjugation to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) and Alexa-Fluor-488 (AF-488) functionalities preserved biological activity, confirmed by (sub)nanomolar EC-values (0.05-0.85 nM) in calcium mobilization assays in transfected CHO-KISS1R cells. Conventional target detection methods using antibodies (Abs) and AF-488-KPs failed to visualize KISS1R in both model (CHO-KISS1R) and native cancer cell lines, likely due to unspecific Abs and rapid KISS1R internalization upon agonist stimulation. However, rapid KISS1R internalization was successfully visualized live-cell imaging using AF-488-KP-10 and novel analogue AF-488-KiSS-34. Furthermore, DOTA-KPs were radiolabeled with Lu-177 in high efficiencies (≥95%) and examined in internalization assays, showing highest uptake (4.8%) and internalization rate (45.9%) for [Lu]Lu-DOTA-KiSS-34 in CHO-KISS1R cells compared to its KP-10 analogue (total uptake: 1.3%; internalization rate: 37.6%). Higher uptakes likely derive from faster binding kinetics, improved KISS1R targeting, and/or slower dissociation as evidenced by oil-based kinetics assays showing higher total uptake for [Lu]Lu-DOTA-KiSS-34 (15.3%) compared to KP-10 (3.8%) and KP-54 (4.5%) counterparts after 30 min. Positron emission tomography/computerized tomography (PET/CT) imaging, urine analysis, and all studies indicate that Ga-68/Lu-177-labeled DOTA-KiSS-34 exhibits superior pharmacodynamics, pharmacokinetics, and stability compared to its KP-10 and KP-54 analogues, which are critically suffering from rapid degradation. These results position DOTA-KiSS-34 as a strong structural lead for KISS1R-based radiotheranostics. Nevertheless, the dynamics between KPs and KISS1R need to be further investigated to fully harness the radiotheranostic potential of KISS1R for TNBC and other cancers.
SPECT Imaging with Bifunctional Technetium-99m Nitrido Bis(thiosemicarbazonato) Complexes Tethered to Inhibitors of the Prostate-Specific Membrane Antigen.
Maclean RC, Fletcher NL, Chu W … +3 more , Lim TX, Tuck KL, Paterson BM
Mol Pharm · 2026 Apr · PMID 41774105 · Publisher ↗
We report the design and synthesis of new conjugates (H) consisting of a bifunctional bis(thiosemicarbazone) chelator conjugated to prostate-specific membrane antigen pharmacophores. Radiolabeling H with the Tc nitrido c...
We report the design and synthesis of new conjugates (H) consisting of a bifunctional bis(thiosemicarbazone) chelator conjugated to prostate-specific membrane antigen pharmacophores. Radiolabeling H with the Tc nitrido core in a one-pot synthesis at 95 °C for 10 min produced the complexes [Tc][TcN()] with high radiochemical purity (RCP > 95%) and excellent stability. evaluation of the [Tc]Tc PSMA complexes demonstrated high cell-surface receptor binding and internalization in cancer cells expressing PSMA (PSMA+), which was inhibited by an excess of a PSMA-specific inhibitor. The most lipophilic of the complexes, [Tc][TcN()], also showed significant nonspecific uptake in the control cell line (PSMA-). SPECT-CT imaging of the four complexes, at 1 and 4 h postinjection and biodistribution studies at 5 h postinjection in BALB/c nude mice with both PSMA+ and PSMA- tumors, demonstrated significant differences in the receptor-specific tumor uptake and clearance pathways. [Tc][TcN()] and the bivalent [Tc][TcN()] showed predominantly renal clearance and the highest PSMA+ tumor uptake and retention. Conversely, [Tc][TcN()] and [Tc][TcN()] displayed gastrointestinal clearance and lower PSMA+ tumor uptake. The work demonstrates that bis(thiosemicarbazones) are effective bifunctional chelators for the development of Tc radiopharmaceuticals targeting PSMA with the potential to be easily modified to target other diseases.
Enhanced Efficacy and Safety of Lu-Anti-CD25 Radioimmunotherapy by Combination with Targeted Anticancer Agents.
Kim JL, Kang CM, Jung KH … +4 more , Kim M, Kim G, Lee H, Lee KH
Mol Pharm · 2026 Apr · PMID 41773645 · Publisher ↗
Anaplastic large cell lymphoma (ALCL) is an aggressive T-cell malignancy for which improved therapeutic strategies are urgently needed. CD25, the α-chain of the interleukin-2 receptor, is abundantly expressed on malignan...
Anaplastic large cell lymphoma (ALCL) is an aggressive T-cell malignancy for which improved therapeutic strategies are urgently needed. CD25, the α-chain of the interleukin-2 receptor, is abundantly expressed on malignant T cells and represents a promising target for antibody (Ab)-based radiotherapeutics. We previously developed a cysteine site-specifically labeled Lu-CD25 Ab that induced complete regression of SUDHL1 tumors but caused dose-limiting bone-marrow suppression at high activities. In the present study, we investigated whether combining reduced-dose Lu-CD25 Ab radioimmunotherapy (RIT) with molecularly targeted inhibitors of ALK (crizotinib) or mTOR (everolimus) could enhance antitumor efficacy while minimizing systemic toxicity. In SUDHL1 cells, crizotinib markedly suppressed ALK phosphorylation, while everolimus potently inhibited phosphorylation of p70S6K, confirming effective pathway blockade. Furthermore, both agents dose-dependently suppressed SUDHL1 cell survival. In SUDHL1 xenografts, treatment with low dose (4.625 MBq) Lu-CD25 Ab alone induced tumor apoptosis and reduced p70S6K activation, while combination therapy with either crizotinib or everolimus further increased cleaved PARP levels, indicating enhanced apoptosis. Longitudinal tumor-growth analysis demonstrated that low-dose Lu-CD25 Ab combined with crizotinib or everolimus produced significantly greater tumor-growth suppression than monotherapies, yielding the lowest tumor burdens and smallest area-under-the-curve values. Kaplan-Meier analysis confirmed prolonged survival in all RIT groups, with the strongest benefit observed in the combination cohorts. Importantly, hematologic, hepatic, and renal parameters remained within normal ranges following combination therapy, reflecting the favorable safety profile of reduced-dose Lu-CD25 Ab. These findings demonstrate that targeted inhibition of ALK or mTOR synergizes with CD25-directed Lu RIT to enhance therapeutic efficacy without increasing toxicity. This combinatorial approach enables radiation-dose reduction while preserving antitumor potency, supporting further preclinical and translational development of Lu-CD25-based combination RIT for ALCL and other CD25-expressing malignancies.
Experimental Evaluation and xDLVO Modeling of Excipient Effects on Protein-Protein Interactions for High-Concentration Formulations.
Ge K, Shen J, Ji Y
Mol Pharm · 2026 Apr · PMID 41770071 · Publisher ↗
The rational design of high-concentration protein formulations remains a significant challenge in biopharmaceutical development, which requires the screening of effective excipients to suppress aggregation. This study pr...
The rational design of high-concentration protein formulations remains a significant challenge in biopharmaceutical development, which requires the screening of effective excipients to suppress aggregation. This study presents a combined experimental and modeling approach to systematically evaluate the effects of key excipients (salts, sugars, polyols, and amino acids) on protein interactions through the measurements of the osmotic second virial coefficient () of bovine serum albumin (BSA) using static light scattering. The results demonstrate that the type and concentration of excipient significantly modulate intermolecular interactions, with repulsive interactions generally enhanced by sugars, polyols, and amino acids, while electrostatic shielding by salts reduces . The exclusion volume (dominant)-water competition (secondary) mechanism was proposed to explain the distinct stabilizing behaviors of the sugars and polyols. In addition, the xDLVO model accurately captures the trends in as a function of excipient type and concentration. This framework provides a powerful predictive method for the early stage screening of excipients, facilitating the development of a stable high-concentration protein formulation.
Attenuated Toxicity and Antitoxic Mechanism via Sodium Iodide Symporter Inhibition-Based Tumor-Selective Delivery in Astatine-211 Radioimmunotherapy.
Takashima H, Tsumura R, Koga Y … +12 more , Anzai T, Yin X, Sato N, Shigekawa Y, Kanayama Y, Nambu A, Usuda S, Haba H, Sakashita S, Inaki A, Manabe S, Yasunaga M
Mol Pharm · 2026 Apr · PMID 41764644 · Full text
Astatine-211 (At) is a promising alpha emitter for cancer treatment, wherein tumor-selective accumulation is pivotal due to its short path length. While sodium ascorbate (SA) successfully protects radioactive antibodies...
Astatine-211 (At) is a promising alpha emitter for cancer treatment, wherein tumor-selective accumulation is pivotal due to its short path length. While sodium ascorbate (SA) successfully protects radioactive antibodies from reactive oxygen species (ROS)-induced denaturation, it does not reduce At distribution in normal organs or mitigate body weight loss. Here, we aimed to attenuate this normal organ uptake. We demonstrated that sodium perchlorate (SP), a competitive inhibitor of the sodium iodide symporter (NIS) expressed in thyroid and gastric mucosal cells, significantly reduced At uptake in the stomach and thyroid in At-radioimmunotherapy (RIT) under SA protection. This favorable biodistribution resulted in significantly milder body weight loss without attenuating the antitumor effect. The combined strategy proved feasible, with no renal toxicity and no exacerbation of transient hematotoxicity or hepatotoxicity. Crucially, NIS inhibition significantly reduced DNA double-strand breaks in stomach and thyroid tissues and helped maintain the thyroid's follicular structure. Overall, we demonstrate that combining SA protection to prevent antibody denaturation with competitive NIS inhibition by SP for greater tumor-selective At delivery is feasible, broadens the therapeutic window, and facilitates the clinical application of At-RIT in cancer treatment.
A Doppler/Near-Infrared-Mediated Visual Real-Time Positioning Microbubble-Based Drug Delivery System Integrating Chemotherapy, Photodynamic Therapy, and Photothermal Therapy for the Treatment of Superficial Tumors.
Zhang Z, Fang C, Ma Y … +8 more , Li A, Dai Y, Chen L, Ren J, Duan M, Chen M, Wang L, Wang C
Mol Pharm · 2026 Apr · PMID 41732842 · Publisher ↗
The treatment of superficial tumors is often limited by poor drug targeting and a lack of real-time visualization of drug distribution. To address these challenges, we developed an RGD-modified microbubble system (TF/ICG...
The treatment of superficial tumors is often limited by poor drug targeting and a lack of real-time visualization of drug distribution. To address these challenges, we developed an RGD-modified microbubble system (TF/ICG-MB@RGD) for ultrasound/near-infrared (NIR) dual-modal imaging-guided chemo-photothermal-photodynamic combination therapy. The microbubbles coloaded tegafur (TF), a chemotherapeutic agent, and indocyanine green (ICG), a photothermal and photosensitizing agent, within a lipid shell surrounding a sulfur hexafluoride (SF) core. The surface was functionalized with cyclic RGD peptides to achieve active targeting to tumor vasculature. Upon intravenous administration, the system allowed real-time visualization of accumulation in tumor tissue via Doppler ultrasound and NIR imaging. Based on the imaging feedback, an 808 nm laser was applied to trigger rapid release of TF and ICG, simultaneously inducing chemotherapy, hyperthermia, and reactive oxygen species generation. In a 4T1 murine breast cancer model, this strategy resulted in a tumor inhibition rate of 72.76% with minimal systemic toxicity. This integrated approach provides a promising theranostic platform for precision treatment of superficial tumors.
Eutectic Coamorphous System of Enzalutamide and Acetyl Maltose: A Strategy for Improved Physical Stability and Aqueous Solubility.
Cichocka-Łokuciejewska J, Knapik-Kowalczuk J, Dulski M … +3 more , Greber KE, Sawicki W, Paluch M
Mol Pharm · 2026 Apr · PMID 41729018 · Full text
The low aqueous solubility of crystalline enzalutamide (ENZ) and the limited physical stability of amorphous ENZ present significant formulation challenges. In this study, we explore binary co-amorphous systems of ENZ wi...
The low aqueous solubility of crystalline enzalutamide (ENZ) and the limited physical stability of amorphous ENZ present significant formulation challenges. In this study, we explore binary co-amorphous systems of ENZ with octaacetyl maltose (acMAL), focusing on the system having eutectic concentration (ENZ + 75 wt % acMAL) as a potential strategy to enhance both stability and solubility. Based on differential scanning calorimetry (DSC) studies of crystalline materials, the eutectic point was identified, while analysis of DSC thermograms of co-amorphous systems revealed pronounced deviations in values of glass transition temperature () from Gordon-Taylor predictions, implicating the existence of strong specific intermolecular interactions. FTIR studies confirmed the presence of heteromolecular bonding within the mixtures. Broadband dielectric spectroscopy (BDS) showed that, although acMAL increases ENZ molecular mobility, the eutectic co-amorphous formulation significantly suppresses recrystallization under isothermal conditions ( = 413 K), delaying crystallization onset by over 30 h and limiting crystallinity to ≤2% after 55 h. The eutectic ENZ + acMAL composition exhibited sustained supersaturation in both aqueous and biorelevant media, demonstrating a balanced combination of efficient drug release and superior stabilization against recrystallization. These results confirm that eutectic formation followed by co-amorphization of ENZ with acMAL effectively addresses the dual challenges of limited physical stability and poor aqueous solubility. This approach provides a mechanistically rational and transferable strategy for improving the performance of poorly water-soluble APIs in pharmaceutical formulations.
Curcumin-Loaded Liposomes (hPLipo/Cur) with Liver-Targeting Properties for Efficient NAFLD Treatment by Alleviating Mitochondrial ROS-Mediated Ferroptosis via NRF2 Pathway.
Wang X, Fang R, Zhao T … +13 more , Ding L, Cai M, Zhang D, Xu N, Yang S, Zhao S, Zhang H, Xie X, Zhang Z, Zhang M, Zhang L, Zhuge Y, Xu B
Mol Pharm · 2026 Apr · PMID 41725216 · Publisher ↗
NAFLD is a rising health problem worldwide with unsatisfied therapies. Curcumin has an ameliorative but limited effect on NAFLD due to its low water solubility. In this study, we innovatively establish the high-PC-conten...
NAFLD is a rising health problem worldwide with unsatisfied therapies. Curcumin has an ameliorative but limited effect on NAFLD due to its low water solubility. In this study, we innovatively establish the high-PC-content liposome-loaded curcumin (hPLipo/Cur) with liver-targeting properties for NAFLD therapy. hPLipo/Cur, composed of DSPC, cholesterol, and DSPE-PEG, has better biocompatibility and water solubility and is loaded with curcumin with high efficiency. hPLipo/Cur is superior to curcumin in improving hepatic histology, as evidenced by reducing lipid deposition and macrophage infiltration in steatohepatitis. Mechanistically, hPLipo/Cur reduces NRF2 degradation and promotes the nuclear translocation of NRF2, as well as the expression of downstream antioxidant genes. The activated NRF2 pathway reduces cellular oxidative stress and the generation of mitochondrial ROS, thereby reducing the accumulation of lipid peroxides and inhibiting ferroptosis in steatohepatitis. In conclusion, hPLipo/Cur reduces mitochondrial ROS-mediated ferroptosis by enhancing the NRF2 pathway to alleviate steatohepatitis, providing a promising strategy for NAFLD treatment.
Hypoxia-Responsive Retinoid Liposomes for Tumor Microenvironment-Activated Differentiation and Metastasis Suppression.
Li C, Liu D, Li X … +3 more , Zhu D, Zhang H, Wang Y
Mol Pharm · 2026 Apr · PMID 41725206 · Publisher ↗
Cancer stem-like cells (CSCs) are a key driving factor of tumor heterogeneity, metastasis, and chemoresistance. All-trans retinoic acid (ATRA) shows strong potential for inducing CSC differentiation and reducing stemness...
Cancer stem-like cells (CSCs) are a key driving factor of tumor heterogeneity, metastasis, and chemoresistance. All-trans retinoic acid (ATRA) shows strong potential for inducing CSC differentiation and reducing stemness; however, its clinical translation is limited by the lack of effective targeting and favorable biodistribution under physiological conditions. Here, we report a hypoxia-responsive ATRA-phospholipid conjugate (RAPC) as a delivery and differentiation-priming strategy, enabling the fabrication of ATRA-loaded liposomes (ATRA@Lip) with high drug loading and microenvironment-activated release. RAPC was synthesized via a modular route incorporating an azobenzene-based hypoxia-cleavable linker between ATRA and a phosphatidylcholine analog. By coassembling RAPC with cholesterol and phospholipid, stable liposomes were obtained with a maximal ATRA loading of 17 wt % at 50 mol % RAPC content. Under normoxia, ATRA@Lip exhibited minimal leakage, whereas hypoxic conditions triggered rapid azo bond cleavage and accelerated ATRA release. Functionally, hypoxia-triggered ATRA@Lip promoted CSC differentiation and enhanced the chemosensitivity of breast cancer cells to albumin-bound paclitaxel (Nab-PTX) in vitro. Collectively, this hypoxia-responsive lipid-drug conjugate platform enables functional CSC priming and improved combination chemotherapy, offering a promising strategy for microenvironment-activated modulation of CSC-associated chemoresistance in metastatic breast cancer.
Curcumin-Fullerene Nanoantioxidant Treats Ulcerative Colitis through Antioxidant and Anti-Inflammatory Mechanisms.
Liu Y, Li M, Wu S … +5 more , Xu J, Bai Y, Bai T, Wang J, Lv X
Mol Pharm · 2026 Apr · PMID 41721788 · Publisher ↗
Active compounds derived from traditional Chinese medicine have attracted considerable interest as potential strategies for alleviating inflammation. However, their clinical translation is often impeded by limitations, i...
Active compounds derived from traditional Chinese medicine have attracted considerable interest as potential strategies for alleviating inflammation. However, their clinical translation is often impeded by limitations, including poor bioavailability and unsatisfactory therapeutic efficacy. To overcome these challenges, a novel fullerene-based nanoantioxidant was developed to enable oral administration and achieve site-specific targeting of inflammatory regions, thereby improving the therapeutic performance of curcumin (CUR) in ulcerative colitis. HA-C60 was synthesized through amidation between amino groups on aminated fullerene and carboxyl groups on hyaluronic acid, yielding a carrier with intrinsic anti-inflammatory properties. studies demonstrated that the carrier possessed free radical scavenging activity. After encapsulation within HA-C60, the resulting CUR nanoparticle formulation (CUR@HA-C60) exhibited both targeted and synergistic anti-inflammatory effects. This carrier markedly enhanced CUR stability in the upper gastrointestinal tract and facilitated site-specific drug accumulation at inflamed colonic sites. Anti-inflammatory efficacy was confirmed by reduced levels of nitric oxide, tumor necrosis factor-α, interleukin-1β, myeloperoxidase, and malondialdehyde, alongside increased expression of interleukin-10, superoxide dismutase, and glutathione, thereby mitigating inflammatory responses and oxidative stress (OS). Furthermore, CUR@HA-C60 demonstrated favorable biosafety profiles in murine models and significantly improved CUR bioavailability by suppressing inflammation, reducing OS-related damage, and restoring intestinal epithelial integrity. These findings indicate that CUR@HA-C60 represents a promising oral therapeutic strategy for the treatment of inflammatory diseases.
An Expert's View on the Application of TIM Technology in the Development of Oral Drug Products.
López Mármol Á, Barker R, Koziolek M … +10 more , Schwabe R, Hens B, Sarcevica I, Butler JM, McAllister M, Taylor E, Chiang PC, Stainforth N, Jones E, Batchelor H
Mol Pharm · 2026 Apr · PMID 41719575 · Publisher ↗
The TIM models tiny-TIM and TIM-1 enable a physiologically relevant in vitro simulation of drug product performance in the upper human gastrointestinal tract under various intake conditions. These systems are considered...
The TIM models tiny-TIM and TIM-1 enable a physiologically relevant in vitro simulation of drug product performance in the upper human gastrointestinal tract under various intake conditions. These systems are considered to be among the most biorelevant oral in vitro models commercially available on the market. This White Paper has been written by a group of experienced users of the TIM systems and shall provide an independent and comprehensive review of the various applications of these systems in the field of pharmaceutical development, as well as current limitations of the systems and how to address them. Recent improvements in the function of the TIM systems, as well as in the use of the data generated, have expanded the application of TIM systems beyond the prediction of food effects and the support of formulation development. For instance, the incorporation of intraluminal concentration and bioaccessibility profiles into in silico models can significantly improve clinical PK predictions and thereby inform decision-making in drug product development in many ways. This work provides an overview of the TIM systems in preclinical and clinical drug product development, supported by selected case studies of various applications. Furthermore, regulatory aspects, the contribution to reducing animal experimentation (3R), and options for future improvement of the TIM systems are highlighted.
Thermal Stability of IgG4 Monoclonal Antibodies: Ambiguous Roles of Cosolutes, Excipients, and Salts in Conformational and Colloidal States.
Yang JY, Burkert O, Mizaikoff B … +1 more , Smiatek J
Mol Pharm · 2026 Apr · PMID 41719208 · Publisher ↗
Peptide and antibody therapeutics are increasingly central to modern drug development, yet their formulation remains challenged by intrinsic instabilities and aggregation tendencies. In this work, we study the thermal st...
Peptide and antibody therapeutics are increasingly central to modern drug development, yet their formulation remains challenged by intrinsic instabilities and aggregation tendencies. In this work, we study the thermal stability of an IgG4 monoclonal antibody (mAb) in the presence of l-arginine and NaCl by combining experimental analysis with a Kirkwood-Buff theoretical framework. The antibody exhibits multistate unfolding transitions, allowing us to disentangle cosolute effects on both conformational and colloidal stability. Our results reveal a dual and concentration-dependent role of cosolutes. Accordingly, l-arginine and NaCl lower unfolding temperatures, thereby destabilizing native domains while simultaneously increasing aggregation temperatures, consistent with aggregation suppression. Contour plot analyses across protein concentrations demonstrate that aggregation inhibition is largely driven by preferential binding at the single-chain level, whereas unfolding destabilization is relatively insensitive to the protein concentration. These findings highlight the synergistic yet ambiguous influence of cosolutes or excipients such that they act as effective aggregation suppressors but promote unfolding of individual domains and vice versa. From a pharmaceutical perspective, this duality complicates rational formulation design. While cosolutes such as l-arginine enhance solubility and colloidal stability, their destabilizing impact on conformational integrity poses risks for therapeutic functionality. Our study underscores the need for careful optimization of cosolute concentrations in antibody formulations, balancing aggregation prevention with preservation of native structure. These insights provide a mechanistic basis for improved formulation strategies for the development of stable biopharmaceuticals.
A Smart Dicyano-Based Fluorescent Probe for In Vivo Amyloid-β Visualization to Illuminate Alzheimer's Disease.
Li X, Yang H, Liang Z … +7 more , Zeng A, Zou Y, Dai J, Zhu B, Yang J, Zhang L, Wang J
Mol Pharm · 2026 Apr · PMID 41716137 · Publisher ↗
Amyloid-β (Aβ) is a key biomarker for Alzheimer's disease (AD), with its abnormal aggregation closely linked to AD pathology. Fluorescence imaging shows promise for Aβ detection but faces challenges such as optical insta...
Amyloid-β (Aβ) is a key biomarker for Alzheimer's disease (AD), with its abnormal aggregation closely linked to AD pathology. Fluorescence imaging shows promise for Aβ detection but faces challenges such as optical instability, a short blood half-life, and poor blood-brain barrier (BBB) permeability. Additionally, only a few probes could target Aβ oligomers with a higher neurotoxicity. In this study, we designed and synthesized 14 fluorescent probes using the dicyano group as an electron acceptor. Among them, emerged as the most promising probe, with a binding affinity for Aβ aggregates ( = 21 nM) and strong binding to Aβ oligomers. It operates at a free-state emission wavelength of above 700 nm, offering low cytotoxicity, good serum stability, and efficient BBB permeability. In brain slice staining, robustly bound to Aβ plaques in both AD mice and human brain slices. Meanwhile, also exhibited the capability of staining Aβ oligomers in the 4 month AD brain slice. Most important, demonstrated robust imaging capabilities in AD mice of different ages and diverse AD mouse models in an in vivo study. These findings suggest that could be a valuable tool for understanding AD pathology and aiding diagnosis.
Evaluation of RBC Targeting As a Strategy to Extend Half-Life and Reduce Nonspecific Tissue Uptake of Biologics.
Cho N, Rafidi H, Morgenstern T … +7 more , Mandikian D, Janezic E, Javier FR, Ho J, Koerber JT, Ferl GZ, Boswell CA
Mol Pharm · 2026 Apr · PMID 41714858 · Publisher ↗
Delivery strategies exploiting red blood cells (RBCs) have been widely pursued, particularly for the purpose of achieving sustained systemic exposure to small molecule drugs. However, limited efforts have been focused on...
Delivery strategies exploiting red blood cells (RBCs) have been widely pursued, particularly for the purpose of achieving sustained systemic exposure to small molecule drugs. However, limited efforts have been focused on applying RBC-inspired delivery strategies to biologic therapies. We aimed to evaluate RBC binding as a potential strategy to achieve systemic half-life extension or reduced tissue accessibility of antibodies, including those having intravascular targets or safety liabilities within peripheral tissues. The effects of RBC targeting on systemic pharmacokinetic (PK) properties was evaluated by measuring exposures of bispecific antibodies targeting a murine RBC surface target, the TER119 antigen, with or without point mutations that diminish binding to the neonatal Fc receptor (FcRn). Additionally, the effects of RBC targeting on specific (tumor) and nonspecific (normal) peripheral tissue uptake of a bispecific antibody targeting the TER119 antigen and a tumor cell surface tumor target (HER2) was assessed by noninvasive imaging and gamma counting. HER2 was used solely as a surrogate peripheral antigen to model potential safety-relevant tissue engagement rather than as a target expected to benefit from RBC binding. Results from an imaging study in tumor bearing mice revealed that RBC binding reduced peripheral uptake in both tumor and normal tissues but increased uptake in the spleen, which acts as both a reservoir for viable RBCs and a site for hemolysis of senescent RBCs. Furthermore, PK and biodistribution studies in normal mice indicated that RBC binding significantly increased the antibody half-life by ∼14-fold in the absence of FcRn-binding. These efforts lay an initial foundation for developing a next generation of biologic therapies with improved PK and therapeutic windows by taking advantage of the unique properties of RBCs.
Homologous Tumor Cell-Derived Delivery of Doxorubicin via Liquid Nitrogen-Treated 4T1 Cells for Targeted Breast Cancer Therapy.
Yang F, Lu Y, Lang X … +7 more , Sun L, Zou H, Zhou Z, Guo Y, Guo Y, Wang X, Han M
Mol Pharm · 2026 Mar · PMID 41711552 · Publisher ↗
Whole cells serve as promising carriers for delivering bioactive molecules owing to their superior biocompatibility, minimal immunogenicity, and extended circulation time. In addition to preserving the intact cellular st...
Whole cells serve as promising carriers for delivering bioactive molecules owing to their superior biocompatibility, minimal immunogenicity, and extended circulation time. In addition to preserving the intact cellular structure and functions, they provide abundant tumor-associated antigens as immune targets, providing a basis for tumor vaccine development. However, conventional chemotherapy with doxorubicin (DOX) suffers from limited tumor-targeting ability and severe systemic toxicity, which markedly restrict its therapeutic efficacy. In this study, we establish a simple liquid nitrogen freezing strategy to load DOX into liquid-nitrogen-treated (LNT) tumor cells for homologous targeting of breast cancer. The optimized formulation achieved a DOX loading of 38.28 ± 1.61 μg/5 × 10 cells. Both in vitro and in vivo studies confirm that the prepared DOX/LNT cell formulation exhibits excellent safety with no proliferative or pathogenic potential. Regarding the antitumor effect of DOX/LNT cells, in vitro cytotoxicity assays reveal an IC value of 1.104 μg/mL, representing a 1.87-fold improvement over free DOX. Furthermore, DOX/LNT cell achieves superior antitumor efficacy by combining chemotherapy with the activation of antitumor immune responses. Following intravenous administration via tail vein injection, in vivo studies demonstrated a tumor inhibition rate of 81.81%, extending the average survival time of breast tumor-bearing mice to 56 days. This approach offers a novel drug delivery system with improved tumor-targeting specificity, representing a precise and effective therapeutic strategy that integrates chemotherapy and immunotherapy for breast cancer treatment.
A Preclinical Study of [Cu]Cu-NOTA-KN035 for Molecular Imaging of PD-L1 in Tumors.
Xu Y, Hua Y, Liu X … +6 more , Shen Q, Meng Q, Shao X, Sheng J, Fu H, Yu C
Mol Pharm · 2026 Mar · PMID 41711141 · Publisher ↗
The programmed cell death protein-1/programmed cell death ligand-1 (PD-1/PD-L1) axis plays a central role in tumor immune regulation, with PD-L1 expression serving as a critical biomarker for patient stratification and r...
The programmed cell death protein-1/programmed cell death ligand-1 (PD-1/PD-L1) axis plays a central role in tumor immune regulation, with PD-L1 expression serving as a critical biomarker for patient stratification and response prediction. Accurate, noninvasive assessment of PD-L1 expression is, therefore, essential for guiding clinical decision-making. KN035 is an ∼79.6 kDa fusion protein comprising a humanized single-domain antibody linked to an Fc fragment, offering a smaller molecular size than conventional monoclonal antibodies. In this study, KN035 was conjugated with p-SCN-Bn-NOTA and radiolabeled with Cu to generate [Cu]Cu-NOTA-KN035 for PET imaging of PD-L1. The tracer showed high radiochemical purity (>95%) and strong binding specificity . PET imaging and biodistribution studies were performed in H1975 (high PD-L1 expression) and A549 (low PD-L1 expression) nonsmall cell lung cancer (NSCLC) xenograft models. Clear tumor visualization was achieved at 4 h postinjection (5.62 ± 0.55%ID/g in H1975; 4.16 ± 0.18%ID/g in A549), with peak uptake at 48 h (12.32 ± 0.66 and 5.72 ± 0.21%ID/g, respectively). Tumor uptake decreased significantly after blocking with excess KN035, confirming the specificity. These results demonstrate the high PD-L1-targeting specificity of [Cu]Cu-NOTA-KN035, suggesting its great potential as a noninvasive diagnostic tool for immunotherapy-based treatments in the future.
Cell-Penetrating Peptides and Supercharged Proteins: A Comprehensive Protocol from Isolation to Cellular Uptake.
Beribisky AV, Sarne V, Huber A … +3 more , Hengstschläger M, Laccone F, Steinkellner H
Mol Pharm · 2026 Mar · PMID 41709632 · Full text
Cell-penetrating peptides (CPPs) and supercharged proteins (SPs) enable efficient intracellular delivery of macromolecules, with expanding applications in basic research and in therapeutic development. Despite their pote...
Cell-penetrating peptides (CPPs) and supercharged proteins (SPs) enable efficient intracellular delivery of macromolecules, with expanding applications in basic research and in therapeutic development. Despite their potential, reproducible workflows for isolation, biochemical characterization, and quantitative uptake analysis remain limited. Here, we present a comprehensive and replicable protocol for the isolation, characterization, and cellular uptake analysis of CPP-fusion proteins (CPP-FPs) and SPs using methyl-CpG-binding protein 2 (MeCP2) constructs as a proof-of-principle model. This workflow combines native protein purification with dynamic light scattering (DLS)-based buffer optimization. Cellular uptake is then assessed and quantified under live-cell conditions using high-content imaging and imaging flow cytometry, with additional assays to probe endocytic trafficking routes, identify CPP-like motifs in SPs, and validate transducing CPP-FP/SP functionality. The protein isolation and DLS-guided buffer screen yield samples with long-term stability. Live-cell fluorescence microscopy and imaging flow cytometry enable discrimination between membrane-bound and internalized signal, providing higher accuracy compared to plate-based readouts. MeCP2 sequence probing has revealed the presence of a CPP-like motif that is critical to its internalization. Finally, validation assays clearly demonstrated CPP-FP/SP activity. This protocol integrates advances in protein biochemistry, structural analysis, and live-cell imaging into a reproducible pipeline adaptable to a wide range of CPP- and SP-based protein constructs and provides a practical framework for downstream mechanistic and therapeutic interventions.
Development of a Novel Approach for Tumor Hypoxia PET Imaging with Nitroreductase-Specific Responsive Radiotracers.
Huang L, Peng S, Liu H … +7 more , Zeng X, Fang J, Wang Y, Zhu H, Zhuang R, Guo Z, Zhang X
Mol Pharm · 2026 Mar · PMID 41705783 · Publisher ↗
Tumor hypoxia is a major contributor to therapeutic resistance, including radio-, chemo-, and immunotherapy, and is closely linked to unfavorable clinical outcomes. PET imaging has shown promise in guiding the clinical t...
Tumor hypoxia is a major contributor to therapeutic resistance, including radio-, chemo-, and immunotherapy, and is closely linked to unfavorable clinical outcomes. PET imaging has shown promise in guiding the clinical treatment of hypoxic tumors. Here, we developed an NTR-responsive PET strategy for tumor hypoxia imaging using radiolabeled nitrogen mustard analogues (F-NTRP and F-NCRP). Cell uptake studies with A549 cells showed that F-NTRP and F-NCRP exhibited approximately two fold higher uptake and good retention under hypoxic conditions. MicroPET imaging of A549 tumor-bearing mice revealed pronounced tumor accumulation of both F-NTRP and F-NCRP, with uptake values of 1.95 ± 0.37 and 2.86 ± 0.49 %ID/g, respectively, and a rapid clearance rate in normal organs. At 45 min postinjection, the tumor/muscle (T/M) ratios of F-NTRP and F-NCRP were 2.62 and 3.44, respectively, both substantially higher than that of F-FMISO (1.40, < 0.05). Following dicoumarin-mediated inhibition of tumor NTR, the uptake and T/M ratio of F-NCRP were decreased to 1.25 ± 0.32 %ID/g and 1.26 ( < 0.001), respectively. Moreover, T/M ratios correlated well with HIF-1α (r = 0.71) and NTR (r = 0.66) levels in the A549 tumor. Collectively, PET imaging demonstrated that F-NCRP specifically targets tumor NTR and can potentially discriminate between varying degrees of hypoxia.
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