Mol. Pharm. [JOURNAL]
Sun
200 papers
Effect of Amorphization on the Punch Sticking Propensity of Organic Powders.
Chandrashekhar Joshi V, Sun CC
Mol Pharm · 2026 May · PMID 42219701 · Publisher ↗
Powder adhesion to tooling surfaces during pharmaceutical tablet manufacturing can compromise product quality and lead to significant production inefficiencies. Although prior studies have assessed adhesive tendencies in...
Powder adhesion to tooling surfaces during pharmaceutical tablet manufacturing can compromise product quality and lead to significant production inefficiencies. Although prior studies have assessed adhesive tendencies in relation to mechanical, particulate, thermal, and surface-chemical properties, these investigations have focused almost exclusively on crystalline materials, leaving the role of amorphization largely unexplored. In this work, using eight chemically diverse model compounds, we systematically investigate how amorphization influences the adhesion of organic powders to steel tooling under realistic compaction conditions and elucidate the underlying mechanisms. Gravimetric measurements for eight cases show that, relative to the crystalline form, amorphization either significantly reduces adhesion (six cases) or produces no measurable change (two cases). Particle morphology does not consistently account for these trends, whereas plasticity analysis reveals a shift in the dominating adhesion mode from contact area-controlled adhesion to contact strength-controlled adhesion. At the molecular level, X-ray photoelectron spectroscopy indicates that amorphization alters the distribution of surface-exposed functional groups that interact with the oxygen-terminated iron oxide layer on steel tooling, leading to differences in adhesion to punch tips. Collectively, these results establish the important role of amorphization-induced changes in surface chemistry in governing powder-tool adhesion and provide a mechanistic framework for understanding adhesion between organic powders and metal surfaces during tablet compression.
A Multitechnique Spectroscopy Platform for Monitoring Heterogeneities in the Higher-Order Structure of mAb Therapeutics.
Agrawal D, Joshi S, Nikita S … +2 more , Sriwastaw S, Rathore AS
Mol Pharm · 2026 May · PMID 42216916 · Publisher ↗
Comprehensive characterization of higher-order structure (HOS) is a regulatory prerequisite for monoclonal antibody (mAb) therapeutics, particularly in establishing biosimilarity. A significant analytical challenge lies...
Comprehensive characterization of higher-order structure (HOS) is a regulatory prerequisite for monoclonal antibody (mAb) therapeutics, particularly in establishing biosimilarity. A significant analytical challenge lies in detecting the early onset of structural heterogeneity within bulk drug substances, where signal dilution often masks precursor states of degradation. This study proposes a novel, multitechnique spectroscopy platform consisting of UV-vis, Far-UV circular dichroism, intrinsic and extrinsic fluorescence, and Fourier transform infrared (FTIR) spectroscopy coupled with chemometrics for monitoring heterogeneities in HOS during the thermal degradation of trastuzumab, an IgG1mAb therapeutic. While size exclusion chromatography (SEC) confirmed a progressive, monotonic accumulation of soluble aggregates, the proposed multitechnique spectroscopic platform yielded richer information. Intrinsic and extrinsic fluorescence identified an early conformational drift characterized by tertiary core destabilization and hydrophobic exposure immediately upon thermal stress (<1 h), well before the onset of global backbone unfolding detected by circular dichroism. Data fusion via principal component analysis (PCA), biplot, and dendrogram allowed for a more nuanced understanding of the HOS evolution through the thermal stress timeline. The data-fusion approach explained in this study offers a robust, high-resolution framework for monitoring structural consistency through direct HOS mapping, offering a powerful solution for monitoring the quality of biotherapeutic products during stability studies as well as for biosimilarity assessments.
Laminin Gamma 2 (LAMC2)-Targeting F(ab') for Zr Immuno-PET Imaging of Human NSCLC Xenograft Models.
Lu R, Zhang J, Bao G … +3 more , Kong Y, Xv D, Wang S
Mol Pharm · 2026 May · PMID 42212402 · Publisher ↗
Laminin gamma 2 (LAMC2) is upregulated in various tumors and is strongly associated with tumorigenesis, aggressiveness, metastasis, and a poor prognosis. This study evaluated the feasibility of Zr-labeled F(ab') fragment...
Laminin gamma 2 (LAMC2) is upregulated in various tumors and is strongly associated with tumorigenesis, aggressiveness, metastasis, and a poor prognosis. This study evaluated the feasibility of Zr-labeled F(ab') fragments of an anti-LAMC2 monoclonal antibody (αLAMC2) for the noninvasive PET/CT imaging of LAMC2-positive nonsmall cell lung cancer (NSCLC) xenografts. LAMC2 expression in NSCLC cell lines was assessed with immunofluorescence staining and Western blotting, and NCIH292 (LAMC2-high) and A549 (LAMC2-low) cells were selected for further analyses. The F(ab') fragment of αLAMC2 was radiolabeled with Zr and evaluated in vitro for radiochemical purity, stability, cellular uptake, and internalization. PET/CT imaging and biodistribution analyses were conducted in NSCLC xenograft models to determine probe uptake and tumor targeting, with Zr-DFO-IgG-F(ab') used as a nontargeted control. The radiochemical purity of the Zr-DFO-αLAMC2-F(ab') probe started at 99.55 ± 0.37% and decreased to 98.00 ± 0.99% in PBS and 96.68 ± 1.66% in 5% FBS by day 5. Compared with A549 cells, NCIH292 cells exhibited substantial probe internalization and uptake at 24 h and 37 °C. Probe uptake was consistently higher in NCIH292 cells than in A549 cells, with NCIH292 exhibiting greater uptake than the control Zr-DFO-IgG-F(ab') at all time points, whereas A549 showed higher uptake than the control only at 24 h. PET imaging revealed significantly greater uptake in LAMC2-high xenografts (NCIH292) than in LAMC2-low xenografts (A549). Biodistribution studies corroborated these findings, showing high accumulation in NCIH292 xenografts (30.66 ± 3.72%ID/g at 72 h) versus low uptake in A549 xenografts (19.06 ± 3.03%ID/g at 72 h). The control probe Zr-DFO-IgG-F(ab')2 exhibited consistently low uptake in both PET imaging and biodistribution analyses. Immuno-PET with Zr-DFO-αLAMC2-F(ab') enabled the robust and specific visualization of LAMC2-expressing xenografts, supporting its potential as a noninvasive molecular imaging probe for the in vivo detection and quantification of LAMC2-positive tumors.
Soluble-Antigen-Based PET/MR for the Specific Detection of BCMA CAR+ T-Cell Lymphoma.
Zhou W, Meng Y, Yan Y … +8 more , Tang L, Yan J, Pan D, Chen C, Xu Y, Wang L, Wang X, Yang M
Mol Pharm · 2026 May · PMID 42207978 · Publisher ↗
Chimeric antigen receptor (CAR) T-cell therapy carries a rare but critical risk of secondary CAR+ T-Cell Lymphoma (CAR-TCL), highlighting the urgent need for precise diagnostic tools. Current noninvasive imaging methods...
Chimeric antigen receptor (CAR) T-cell therapy carries a rare but critical risk of secondary CAR+ T-Cell Lymphoma (CAR-TCL), highlighting the urgent need for precise diagnostic tools. Current noninvasive imaging methods struggle to differentiate CAR-TCL from inflammatory sequelae, while biopsy presents several clinical challenges. In this study, we developed a novel positron emission tomography (PET) imaging probe, [Ga]Ga-BCMA-NOTA, by repurposing the extracellular domain of B-cell maturation antigen (BCMA-ECD) as a specific molecular recognizer for the BCMA-CAR single-chain variable fragment (scFv). Leveraging the high-affinity antigen-antibody interaction, the recombinant BCMA probe was produced with high purity (>98%) and demonstrated nanomolar affinity for the BCMA CAR scFv ( = 5.17 nM). The probe was synthesized with excellent radiochemical purity (>95%) and a good radiochemical yield (>50%). In preclinical xenograft models, [Ga]Ga-BCMA-NOTA PET/MR imaging demonstrated high and specific uptake in BCMA CAR-positive tumors (2.56 ± 0.61%ID/g), compared to negligible uptake in BCMA CAR-negative tumors (0.21 ± 0.05%ID/g), with a tumor-to-muscle (T/M) ratio of 16.0 ± 3.8. The uptake was blockable, confirming the probe's target specificity in vivo. Importantly, in a head-to-head comparison, [Ga]Ga-BCMA-NOTA provided significantly superior imaging contrast and a stronger correlation with tumor burden ( = 0.93) compared to [F]FDG ( = 0.76). This BCMA CAR scFv-specific probe offers a powerful new tool for the precise noninvasive visualization of BCMA CAR-positive malignancies, with significant clinical translational potential for ensuring the long-term safety and monitoring of CAR-T therapies.
Voices in : Meet Dr. Hannes Steinkellner, Who Is Turning Engineered Proteins into Intracellular Therapies Using Cell-Penetrating Peptides.
Steinkellner H
Mol Pharm · 2026 May · PMID 42206860 · Publisher ↗
Abstract loading — click title to view on PubMed.
Insights into the Structural Coexistence of Hexameric States in Microcrystalline Insulin Formulations in the Solid State.
Rohilla KK, Bhardwaj Y, Pandey MK
Mol Pharm · 2026 May · PMID 42206399 · Publisher ↗
The structural complexity and insoluble nature of solid-state therapeutic proteins pose major challenges for their pharmaceutical quality control. Many conventional analytical methods cannot be applied directly to fully...
The structural complexity and insoluble nature of solid-state therapeutic proteins pose major challenges for their pharmaceutical quality control. Many conventional analytical methods cannot be applied directly to fully formulated drug products, leaving important aspects of their higher-order structure (HOS) poorly understood. In this work, we use high-resolution, proton-detected solid-state NMR (ssNMR) spectroscopy under fast magic-angle spinning (MAS) to probe the atomic-level HOS of insulin directly in its native formulated state. Recombinant human insulin powder was compared with three commercial Neutral Protamine Hagedorn (NPH) crystalline suspensions: Humulin 70/30, Insugen 70/30, and NovoMix 30. Two-dimensional (2D) C-H correlation spectra showed that the overall insulin fold is conserved across all samples. In contrast, the 2D N-H spectra provided a much more sensitive fingerprint of HOS and revealed clear structural differences. The amorphous insulin powder exhibits a conformationally heterogeneous ensemble characterized by a complex interplay of static disorder, dynamic averaging, and potential partial unfolding reflecting a nonhexameric, T-like architectural state. In contrast, the formulated suspensions show significantly increased N chemical shift dispersion, consistent with more rigid, hexameric assemblies. Inspection of the allosteric reporter residues Gly and Thr shows that the NPH crystals are not locked into a single R conformation. Instead, multiple allosteric states are present, plausibly reflecting differences in hydration and phenolic ligand binding within the lattice. The NovoMix 30 formulation (insulin aspart) exhibits further line broadening, consistent with increased lattice disorder introduced by the Pro → Asp substitution. These observations illustrate how solid-state NMR can resolve formulation-dependent conformational heterogeneity and provide structural support for biosimilarity evaluation in solid-state protein therapeutics.
Assays for Measuring the Cell Permeability of Proteolysis-Targeting Chimeras (PROTACs): Performance, Correlations, Applicability and Recommendations.
Brenner C, Cui Y, Schmidt D … +3 more , Kirchmair J, Tan L, Braun N
Mol Pharm · 2026 May · PMID 42203198 · Publisher ↗
Most Proteolysis-Targeting Chimeras (PROTACs) are beyond-Rule-of-Five compounds, and optimizing their ability to cross cell membranes is a central topic in PROTAC therapeutic research. In this study, we systematically in...
Most Proteolysis-Targeting Chimeras (PROTACs) are beyond-Rule-of-Five compounds, and optimizing their ability to cross cell membranes is a central topic in PROTAC therapeutic research. In this study, we systematically investigate the performance and assay correlations of PROTAC cell permeability assays and the chemical space in which these assays provide reliable results. Our work builds on a comprehensive data set of 3271 PROTACs spanning diverse chemical and target spaces, providing unprecedented coverage of PROTAC cell permeability data. These compounds were measured in up to three different types of assays: the NanoBRET E3 ligase target engagement assay, the Caco-2 assay, and a P-glycoprotein (P-gp) efflux liability assay. We find a strong correlation between readouts from the NanoBRET E3 ligase target engagement assay (availability shift) and those from the Caco-2 assay (apparent intrinsic permeability, basolateral-to-apical permeability, and efflux ratio), while P-gp liability derived from the P-gp assay cannot easily be translated to the Caco-2 efflux ratio. In the process, we also investigate the impact of physicochemical properties included in the Rule-of-Five on the applicability of the assays. Based on these findings, we derive guidelines and data-driven recommendations for the use of cell permeability assays in PROTAC optimization. While this study focuses on PROTACs, some of the observations may be relevant to other modalities with beyond-Rule-of-Five physicochemical properties.
Linker Optimization in Lu-177 Labeled αvβ6-Integrin Binding Peptide Trimers for Targeted Radionuclide Therapy of Cancer.
Nguyen NT, Rheinfrank T, Stangl S … +3 more , Reissig F, Kossatz S, Notni J
Mol Pharm · 2026 May · PMID 42200221 · Publisher ↗
Theranostic radiopharmaceuticals exploit the same molecular target for diagnosis and therapy. Among emerging pan-cancer targets, αvβ6-integrin is highly expressed on various malignant cell types and can be imaged clinica...
Theranostic radiopharmaceuticals exploit the same molecular target for diagnosis and therapy. Among emerging pan-cancer targets, αvβ6-integrin is highly expressed on various malignant cell types and can be imaged clinically with Ga-68-Trivehexin. However, therapeutic αvβ6-integrin-directed radioligands remain scarce. Here, Lu-177-labeled multimerics of the cyclic nonapeptides Tyr2, sequence c(YRGDLAYp(Me)K), were constructed using the tetrafunctional chelator DOTPI by means of CuAAC-based and evaluated as αvβ6-integrin-targeted radiotherapeutics. PEG linkers of increasing length (PEG0, PEG3, PEG7, PEG11) were introduced between the chelator and peptides to modulate pharmacokinetics and cellular processing. The Lu-177-labeled conjugates displayed comparable polarity (log ≈ -2) and αvβ6-integrin affinities determined by ELISA (IC ∼ 0.2-0.5 nM), indicating minimal effects of PEG length on in vitro parameters. In αvβ6-positive H2009 cells, PEG linkers markedly enhanced and prolonged receptor-mediated uptake and internalization, consistent with improved multivalent engagement. In H2009 xenograft mice, PEG linkers reduced early blood-pool activity and increased tumor uptake at 24 h, while gelofusine (4% succinylated gelatin in Ringer's acetate) efficiently mitigated the elevated renal retention (up to 92% reduction). Performance gains plateaued beyond PEG7, identifying the PEG7 trimer () as the preferred lead due to favorable tumor uptake/retention. exhibited high selectivity (65- to 671-fold) over other integrin subtypes, αvβ6-integrin-dependent radiotoxicity to tumor cells, and sustained tumor retention for up to 6 days according to μSPECT.
Intranasal Administration of a Neuromedin-U Receptor 2-Specific Peptide Agonist for Obesity Treatment: Effect of the Stability on Brain Delivery.
Tanaka A, Yamashita A, Takayama K … +3 more , Hayashi Y, Katsumi H, Furubayashi T
Mol Pharm · 2026 May · PMID 42185239 · Publisher ↗
Neuromedin U (NMU) is a neuropeptide that was first identified in porcine spinal cord and exhibits diverse biological activities. It serves as an endogenous ligand for the human Neuromedin U receptor 2 (NMUR2), which is...
Neuromedin U (NMU) is a neuropeptide that was first identified in porcine spinal cord and exhibits diverse biological activities. It serves as an endogenous ligand for the human Neuromedin U receptor 2 (NMUR2), which is expressed exclusively in the hypothalamus. Because NMUR2 is involved in the regulation of appetite suppression and energy metabolism, it is considered a promising target for obesity treatment. CPN is a medium-sized peptide with selective agonistic activity against NMUR2. In this study, we evaluated the effect of peptide stability on brain delivery and the pharmacological effects after intranasal administration of two CPNs (CPN-116 and CPN-219) with different in vitro stabilities. The results of in vitro stability studies showed that both CPNs were more stable in the cerebrospinal fluid (CSF) than in the plasma and that the stability of CPN-219 in the serum, CSF, brain, and nasal cavity was better than that of CPN-116. For both CPNs, brain concentrations were higher in the nasal administration group than in the intraperitoneal administration group, and weight gain was also suppressed in the nasal administration group compared to the control group. Furthermore, both brain delivery and suppression of weight gain after nasal administration of CPN-219, which has good in vitro stability, were superior to those of CPN-116. It is evident that the stability of the peptide is an important factor for the direct delivery of peptides into the brain by intranasal administration.
Platelets as Programmable Drug Depots: Toward Predictive and Mechanistic Drug Loading.
Moskalensky AE
Mol Pharm · 2026 May · PMID 42175967 · Publisher ↗
Efficient drug delivery remains a major challenge in pharmaceutical science, with synthetic nanocarriers often facing limitations in real biological systems. While blood platelets have been explored as biomimetic carrier...
Efficient drug delivery remains a major challenge in pharmaceutical science, with synthetic nanocarriers often facing limitations in real biological systems. While blood platelets have been explored as biomimetic carriers, their intrinsic capacity for molecular storage and stimulus-triggered release remains largely underutilized. Here, we propose that platelet granules can be viewed as programmable drug depots, whose loading can be rationally controlled using physicochemical principles and endogenous transport mechanisms. In contrast to empirical approaches, we outline a framework based on two complementary pathways: passive accumulation driven by ion trapping of weakly basic compounds in acidic granules, and active uptake mediated by vesicular transporters. Building on recent advances in structural biology and computational modeling, we further argue that drug loading into platelets can be predicted and optimized using molecular descriptors, docking approaches, and machine learning models to estimate transporter compatibility and accumulation efficiency. This perspective introduces a conceptual design loop linking prediction and molecular optimization─i.e., enabling both selection of suitable drug candidates and rational modification of their structures to enhance platelet compatibility. Together, these concepts position platelets not simply as biomimetic carriers but as model-guided and engineerable platforms for drug delivery, bridging biological systems with rational design strategies.
High-Density Lipoprotein Nanoparticles Delivering Liver X Receptor Agonist for the Treatment of Age-Related Macular Degeneration.
Mei L, Weh E, Yu M … +5 more , Walsh L, Fahim AT, Liu Y, Besirli CG, Schwendeman A
Mol Pharm · 2026 Jul · PMID 42172633 · Publisher ↗
Liver X nuclear receptor (LXR) agonists are promising therapeutic agents whose efficacy has been demonstrated and for the treatment of atherosclerosis. The accumulation of lipids is a common pathogenic mechanism betwee...
Liver X nuclear receptor (LXR) agonists are promising therapeutic agents whose efficacy has been demonstrated and for the treatment of atherosclerosis. The accumulation of lipids is a common pathogenic mechanism between both atherosclerosis and age-related macular degeneration (AMD). As such, a growing number of studies have implicated LXR agonists as a potential therapeutic option for the treatment of AMD. Unfortunately, the clinical application of LXR agonists has been hindered by their undesirable side effects when administered systemically, as well as their limited aqueous solubility, necessitating a drug delivery system (DDS) for targeted local delivery. Synthetic high-density lipoprotein (sHDL) nanoparticles have intrinsic cholesterol transport and anti-inflammatory functions, making them ideal for the treatment of diseases where cholesterol accumulation and inflammation contribute to the pathology. These nanoparticles also excel as an effective DDS for hydrophobic drugs due to their ability to encapsulate drugs in their hydrophobic core. Thus, we developed a new strategy for AMD treatment by encapsulating an LXR agonist, T0, in sHDL nanoparticles (sHDL-T0). Our studies showed that sHDL-T0 can effectively upregulate cholesterol transport from mature iPSC-RPE (retinal pigment epithelium) due to increased transcription of cholesterol transporters and by serving as an acceptor for effluxed cholesterol. Encapsulating T0 in sHDL nanoparticles increased the aqueous solubility of T0 significantly, allowing for direct delivery to the eye without organic solvents. delivery of sHDL-T0 caused upregulation of the Abca1/g1 transporters in the retina and retinal pigment epithelium, suggesting that T0 can escape sHDL nanoparticles and stimulate the LXR pathway. Most importantly, in a murine model of nonexudative (dry) age-related macular degeneration (dAMD), sHDL-T0 administration resulted in the down-regulation of , and mRNA expression, and reduced the number of CD45-positive inflammatory cells in comparison to vehicle-treated eyes. Our results provide proof-of-concept for using sHDL nanoparticles to deliver the LXR agonist T0 to restore cholesterol homeostasis and reduce local inflammation in the outer retina, offering a novel therapeutic strategy for dAMD.
Butyrate Regulates the Blood-Brain Barrier Transport and Intraendothelial Accumulation of Alzheimer's Disease Amyloid-Beta Peptides.
Veerareddy V, Wang Z, Kashyap PC … +1 more , Kandimalla KK
Mol Pharm · 2026 Jul · PMID 42166642 · Publisher ↗
Alzheimer's disease (AD) is characterized by the pathological deposition of amyloid β (Aβ) peptides as amyloid plaques and by cerebrovascular dysfunction, both of which drive AD progression. Butyrate, a gut microbial met...
Alzheimer's disease (AD) is characterized by the pathological deposition of amyloid β (Aβ) peptides as amyloid plaques and by cerebrovascular dysfunction, both of which drive AD progression. Butyrate, a gut microbial metabolite, is reduced in AD patients, and its supplementation has been shown to improve cognition and reduce the amyloid burden in animal models. However, the underlying mechanisms are unclear. Our previous studies demonstrated that insulin signaling regulates Aβ transport kinetics at the blood-brain barrier (BBB). In this study, we investigated whether butyrate reduces intraendothelial Aβ accumulation and improves BBB integrity through modulation of insulin signaling pathways. The effect of butyrate on Aβ42 accumulation was assessed by flow cytometry in polarized BBB endothelial cell culture models. Activation of insulin signaling and expression of key BBB constituents involved in Aβ transport and accumulation [p-glycoprotein (P-gp), the receptor for advanced glycation end products (RAGE)], as well as BBB integrity (tight junction protein, claudin-5), were evaluated by using Western blotting and confocal microscopy. The roles of insulin signaling nodes, including AKT, ERK, mTOR, and p38 phosphorylation, were investigated by using specific inhibitors MK2206, Trametinib, Rapamycin, and VX-745, respectively. In addition, the effect of butyrate on BBB receptors and transporters involved in Aβ trafficking was examined in vivo in mouse brains colonized with butyrate-producing bacteria via immunohistochemistry. Butyrate significantly decreased endothelial Aβ42 accumulation, an effect associated with the activation of insulin signaling, particularly AKT and ERK phosphorylation. Inhibitor studies established the critical role of these pathways, as co-incubation with MK2206 (AKT inhibitor) or Trametinib (ERK inhibitor) reversed the protective effect of butyrate and increased Aβ42 accumulation, whereas inhibition of mTOR or p38 had no significant effect. Moreover, butyrate restored Aβ-induced reductions in the P-gp efflux transporter expression and claudin-5 tight junction protein levels. These findings were supported by in vivo studies demonstrating upregulation of the tissue inhibitor of metalloproteinases-2 (TIMP-2), a protein associated with AKT activation and extracellular matrix stabilization in mice colonized with butyrate-producing bacteria. In conclusion, our data demonstrate that butyrate reduces Aβ42 uptake at the BBB endothelium by activating the AKT and ERK arms of the insulin signaling pathway. These changes may also enhance BBB integrity by increasing claudin-5 expression, stabilizing the extracellular matrix, and upregulating TIMP-2. Collectively, this study highlights butyrate as a potential therapeutic modulator of AD-associated BBB dysfunction.
Properties of Drug Linkers Affecting Hydrophobic Interaction Chromatography for Determining the Drug-to-Antibody Ratio of Antibody-Drug Conjugates.
Kusama T, Mimura M, Nakae R … +4 more , Ng SY, Shigemoto R, Hirakura Y, Kawasaki N
Mol Pharm · 2026 Jul · PMID 42166563 · Publisher ↗
Antibody-drug conjugates (ADCs) are a fascinating modality in cancer therapy that combine the specificity of monoclonal antibodies (mAbs) with the cytotoxicity of potent drugs. To ensure the product quality of ADCs, the...
Antibody-drug conjugates (ADCs) are a fascinating modality in cancer therapy that combine the specificity of monoclonal antibodies (mAbs) with the cytotoxicity of potent drugs. To ensure the product quality of ADCs, the drug-to-antibody ratio (DAR), a critical quality attribute (CQA), must be evaluated at quality control (QC) testing. Hydrophobic interaction chromatography (HIC) is considered the gold standard for DAR determination due to advantages such as the ability to evaluate drug load distribution under native conditions. For some ADCs, however, DAR cannot be determined by HIC analysis due to poor peak separation of DAR variants, but the cause of this poor peak separation is unclear. In this study, we investigated which ADC properties impact the peak separation of DAR variants in HIC. HIC analyses of ADCs with different mAb and drug linker (DL) combinations indicated that peak separation was impacted by DL properties rather than mAb properties. A systematic comparison using model ADCs prepared with trastuzumab and various drug linkers revealed that the linker length is a critical structural determinant of peak separation. We hypothesized that a linker that was too short restricted the steric accessibility of drug moieties on ADCs, resulting in weak retention on the stationary phase of the HIC column. Accordingly, we investigated the correlation between linker length and steric accessibility using an enzyme-linked immunosorbent assay (ELISA) to evaluate the binding affinity of biotin, which is conjugated to ADCs as a model drug, for streptavidin. The results supported the notion that short linkers are less preferable for the steric accessibility of the drug moieties. Our findings will aid in understanding the mechanism of HIC peak separation and contribute to the selection of appropriate analytical methods based on ADC structures, even within the accelerated development timelines required to supply new drugs to patients in need.
Voices in : Meet Dr. Paul Klauser, Who Is Developing the Next Generation of Radiotheranostic Drugs to Improve Cancer Therapy.
Klauser PC
Mol Pharm · 2026 May · PMID 42161860 · Publisher ↗
Abstract loading — click title to view on PubMed.
Insect-Cell-Produced AAV2 Vectors Activate Myeloid Innate Immune Response Featuring NETosis.
Liu X, Zhou Y, Tang Z … +3 more , Li W, Wei S, Han K
Mol Pharm · 2026 Jun · PMID 42157557 · Publisher ↗
Recombinant adeno-associated virus (rAAV) is a leading platform for gene therapy. However, host immune responses remain a major barrier to its safe and effective clinical application. As insect cell systems have emerged...
Recombinant adeno-associated virus (rAAV) is a leading platform for gene therapy. However, host immune responses remain a major barrier to its safe and effective clinical application. As insect cell systems have emerged as a valuable production platform with unique advantages, we examined the immunogenicity of insect-cell-derived rAAV2. Temporal transcriptomic profiling in mice revealed a rapid and sustained innate immune response dominated by myeloid cell activation. Notably, we observed prominent enrichment of the neutrophil extracellular trap (NETosis) pathway. Functional assays in isolated neutrophils confirmed that rAAV2 directly induces a pro-NETotic response marked by reactive oxygen species (ROS) production and NET formation. Degradation of extracellular DNA via DNase I significantly attenuated hepatic inflammatory signaling and enhanced vector transduction, indicating that NET formation functions as a biological barrier to rAAV2-mediated gene transfer. Our findings identify NETosis as a previously underappreciated immune mechanism triggered by insect-cell-produced AAV vectors, offering new insights into vector immunogenicity and revealing potential targets to improve the safety and efficacy of AAV-based gene therapies.
Establishment of Ga-DOTA-Based Pretargeted Radioimmunodiagnosis.
Veach DR, Burnes Vargas D, Islam B … +17 more , Lee SG, Gajecki L, Vaughn BA, Yang G, Kalidindi TM, Pillarsetty NVK, Salehi N, Jaswal AP, Saha S, Le Roux AB, Xu H, Guo HF, Ouerfelli O, Cheung NV, Krebs S, Larson SM, Cheal SM
Mol Pharm · 2026 Jun · PMID 42151769 · Full text
DOTA-radiometal (DOTA: 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) hapten complexes of β-emitters (Lu or Y) and α-emitters (Ac) can be selectively targeted to tumors using an antitumor/anti-DOTA bispecific...
DOTA-radiometal (DOTA: 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) hapten complexes of β-emitters (Lu or Y) and α-emitters (Ac) can be selectively targeted to tumors using an antitumor/anti-DOTA bispecific antibody (BsAb) approach in pretargeted radioimmunotherapy (DOTA-PRIT). Clinical translation of DOTA-PRIT may be accelerated by the development of a Ga companion diagnostic for PET imaging. Here, we report a novel radiohapten, which uses a 1,4,7-triazacyclononane,1-glutaric acid-4,7-acetic acid (NODAGA) chelator for Ga labeling while retaining Lu in a benzyl-DOTA moiety for high-affinity antibody recognition (NODAGA-Pr; Pr = Proteus). Radiolabeling of NODAGA-Pr with Ga showed high radiochemical yield and purity, and studies confirmed high radiostability and negligible serum-protein binding. , using a DOTA-PRIT system targeting the GPA33 antigen in a human colorectal cancer mouse model, [Ga]Ga-NODAGA-Pr demonstrated efficient tumor uptake and high-contrast PET imaging. Additional studies using an established DOTA-hapten test system─human embryonic kidney 293T cells expressing a transmembrane-anchored anti-DOTA scFv huC825 further confirmed its PRIT performance. These findings support [Ga]Ga-NODAGA-Pr as a promising agent for pretargeted immunoPET and for imaging engineered cells expressing a radiohapten capture reporter gene, with favorable pharmacokinetics and tumor targeting well matched to existing therapeutic DOTA-radiohaptens.
An F-labeled ECL1i-Based C-C Chemokine Receptor Type 2 Radiotracer for Cardiac Inflammation Imaging.
Song S, Ren W, Ding D … +7 more , Ju M, Wang K, Zhou Y, Gu Y, Sun R, Fang W, Wang L
Mol Pharm · 2026 Jun · PMID 42150152 · Publisher ↗
C-C chemokine receptor type 2 (CCR2) monocytes and macrophages play a critical role in cardiovascular inflammation. In this study, we developed an F-labeled CCR2-targeted positron emission tomography (PET) radiotracer, [...
C-C chemokine receptor type 2 (CCR2) monocytes and macrophages play a critical role in cardiovascular inflammation. In this study, we developed an F-labeled CCR2-targeted positron emission tomography (PET) radiotracer, [F]AlF-NOTA-PEG-ECL1i, for the noninvasive imaging of cardiac inflammation. Using a one-step [F]AlF-NOTA chelation strategy, [F]AlF-NOTA-PEG-ECL1i was efficiently synthesized from [F]fluoride and exhibited excellent radiochemical purity (>98%), high in vitro and in vivo stability, and a low lipid-water partition coefficient (log = -4.52 ± 0.19). Biodistribution studies demonstrated predominant renal clearance and low nonspecific background uptake. In mice and swine models of cardiac ischemia-reperfusion (I/R) injury, radiotracer uptake increased in the injured myocardium. This observation was further validated by myocardial perfusion imaging, triphenyltetrazolium chloride staining, and ex vivo autoradiography. Furthermore, longitudinal PET/computed tomography (CT) from baseline to 24 days after I/R injury demonstrated that [F]AlF-NOTA-PEG-ECL1i uptake in the injured myocardium was consistent with time-dependent changes in CCR2 cell density. These results support [F]AlF-NOTA-PEG-ECL1i as a promising CCR2-targeted PET radiotracer for imaging cardiac inflammation.
Construction of a Zr-Labeled Specific Antibody Fragment for the Noninvasive Detection of Mesothelin-Overexpressing Tumors.
Hou X, Yang L, Liu F … +7 more , Wang F, Liu S, Yao Y, Wang Z, Qi C, Zhu H, Yang Z
Mol Pharm · 2026 Jun · PMID 42117406 · Publisher ↗
Numerous clinical trials on mesothelin (MSLN)-targeted antibodies have provided preliminary evidence supporting their clinical translation, highlighting the importance of patient screening. Although anti-MSLN chimeric an...
Numerous clinical trials on mesothelin (MSLN)-targeted antibodies have provided preliminary evidence supporting their clinical translation, highlighting the importance of patient screening. Although anti-MSLN chimeric antigen receptor T (CAR-T) cell therapy has not been approved by the Food and Drug Administration (FDA) for solid tumor treatment yet, its high expression across various malignancies has made it a major research focus. Prescreening patients suitable for anti-MSLN CAR-T cell therapy may boost both treatment efficacy and safety. The ScFv component of anti-MSLN CAR-T cells retains MSLN targeting capability and serves as a suitable molecular precursor for probe development. In this study, the positron-emitting nuclide Zr was utilized to label the antibody fragment MA4 (comprising the ScFv segment of MSLN fused with the human Fc segment), while a mouse monoclonal antibody (Sino Biological, 13128-MM08) served as a control, enabling the development of two immunoPET probes for MSLN imaging. Following purification, Zr-MA4 exhibited high radiochemical purity (>99%) and molar activity (24.2-31.9 GBq/μmol) and maintained >95% radiochemical purity over 6 days in both PBS and 5% human serum albumin (HSA). In the MSLN strongly positive LS174T tumor model, the Zr-MA4 probe demonstrated expedited tissue penetration and tumor accumulation. The immunohistochemical analysis revealed that LS174T tumors exhibited strong positivity for MSLN, whereas the U87MG group demonstrated only mild expression. Dosage estimation studies have determined that the effective dose of Zr-MA4 is 0.142 mSv/MBq, which complies with established safety standards. Overall, both probes exhibit robust in vitro stability and demonstrate feasibility for MSLN-targeted imaging in vivo.
Controlling the Size and Shape of Monoclonal Antibody Particles through Crystallization Process Design.
Codan L, Larpent P, Kalapos-Dobszay R … +4 more , Medrán B, Reichert P, Sirota E, Cote A
Mol Pharm · 2026 Jun · PMID 42117276 · Publisher ↗
The crystallization of monoclonal antibodies is deemed particularly challenging due to their large size and structural flexibility. An additional challenge is represented by the tendency of mAb crystals to rapidly lose t...
The crystallization of monoclonal antibodies is deemed particularly challenging due to their large size and structural flexibility. An additional challenge is represented by the tendency of mAb crystals to rapidly lose their crystallinity when removed from liquid and exposed to air or nitrogen, which complicates filtration. This study aims to develop crystallization design strategies for mAbs using the caffeine cocrystal of pembrolizumab as a model. Optimal crystallization conditions are identified by minimizing cycle times for crystallization and filtration. Metrics are introduced to rank the filtration performance of the different crystalline particle populations. This work focuses on the crystallization of caffeine cocrystal-1 of pembrolizumab, which is consistently generated upon charge of the crystallizing agent poly(ethylene glycol) 3350 (PEG) and of mediating salts, among others, dextran sodium sulfate (DSS). This study also demonstrates that seeding is feasible in the crystallization of mAbs and consistently shortens the cycle times for crystallization and improves the robustness of the process. As a proof of concept, seeded semicontinuous crystallizations are performed to enhance supersaturation control during crystallization when stringent impurity rejection and particle attribute specifications must be met. The isolation of crystalline mAb particles presents considerable challenge, and there are no documented studies that have established precedent for this aspirational goal. Finally, to avoid amorphization of the particles during isolation, a conservative approach is adopted to only partially deliquor the filter cake and to leave enough supernatant to fully submerge the filter cake to prevent air or nitrogen exposure. Assuming multiple filtration steps may be required to meet drug substance specifications, this study demonstrates that the crystal form can be retained through successive filtration and dilution/wash cycles with the supernatant composition adjusted after each filtration. This outcome demonstrates that the particles are resistant to changes in the solvent composition, such that the final product could be swapped from the matrix best suited for crystallization to a matrix more suitable for product delivery.
Correction to "Detection of Voltage-Gated Potassium Channels 1.3 via Immuno-PET Visualizes Rheumatoid Arthritis".
Cheng S, Quan Y, Long X … +4 more , Lan X, Jiang D, Liu K, Fan C
Mol Pharm · 2026 Jun · PMID 42113037 · Publisher ↗
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