Solution biophysics identifies lipid nanoparticle non-sphericity, polydispersity, and dependence on internal ordering for efficacious mRNA delivery DOI Creative Commons
Marshall S. Padilla, Sarah J. Shepherd, Andrew R. Hanna

и другие.

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2024, Номер unknown

Опубликована: Дек. 22, 2024

Abstract Lipid nanoparticles (LNPs) are the most advanced delivery system currently available for RNA therapeutics. Their development has accelerated since success of Patisiran, first siRNA-LNP therapeutic, and mRNA vaccines that emerged during COVID-19 pandemic. Designing LNPs with specific targeting, high potency, minimal side effects is crucial their successful clinical use. These characteristics have been improved through microfluidic platforms, which enhanced efficacy uniformity LNP batches. However, our understanding how composition mixing method influences structural, biophysical, biological properties resulting particles remains limited, hindering LNPs. Our lack structural extends from physical compositional polydispersity LNPs, render traditional characterization methods, such as dynamic light scattering (DLS), unable to accurately quantitate physicochemical In this study, we address challenge structurally characterizing polydisperse formulations using emerging solution-based biophysical methods higher resolution provide data beyond size polydispersity. techniques include sedimentation velocity analytical ultracentrifugation (SV-AUC), field flow fractionation followed by multi-angle (FFF-MALS), size-exclusion chromatography in-line synchrotron small-angle X-ray (SEC-SAXS). Here, show intrinsic in size, loading, shape, these parameters dependent on both formulation technique lipid composition. Lastly, demonstrate can be employed predict transfection three models examining relationship between translation characteristics. We envision employing will essential determining structure-function relationships, facilitating creation new design rules future

Язык: Английский

A perspective on bleb and empty LNP structures DOI

Jens B Simonsen

Journal of Controlled Release, Год журнала: 2024, Номер 373, С. 952 - 961

Опубликована: Авг. 8, 2024

Язык: Английский

Процитировано

19
MicroRNA-targeting nanomedicines for the treatment of intervertebral disc degeneration DOI Creative Commons
Hussein H. Genedy, P. Humbert,

Bilel Laoulaou

и другие.

Advanced Drug Delivery Reviews, Год журнала: 2024, Номер 207, С. 115214 - 115214

Опубликована: Фев. 22, 2024

Язык: Английский

Процитировано

13
Retention mechanism in combined hydrodynamic and slalom chromatography for analyzing large nucleic acid biopolymers relevant to cell and gene therapies DOI Creative Commons
Fabrice Gritti, Kevin D. Wyndham

Journal of Chromatography A, Год журнала: 2024, Номер 1730, С. 465075 - 465075

Опубликована: Июнь 21, 2024

Slalom chromatography (SC) was discovered in 1988 for analyzing double-stranded (ds) DNA. However, its progress impeded by practical issues such as low-purity particles, sample loss, and lack of a clear retention mechanism. With the rise cell gene therapies availability today bio-inert ultra-high-pressure liquid (UHPLC) columns systems, SC has regained interest. In SC, elution order is opposite to that observed hydrodynamic (HDC): larger DNA molecules are more retained than small ones. Yet, underlying mechanism remains elusive. We provide physicochemical background necessary explain, at microscopic scale, full transition from HDC This includes persistence length macromolecule (representing stiffness), their relaxation time (τR) non-equilibrium contour equilibrium entropic configuration, relationship between mobile phase shear rate (〈γ̇〉) packed extended length. propose relevant model account simultaneous impact (HDC) on factors series large linear dsDNAs (ranging 2 48 kbp). data were acquired using MaxPeakTM Columns with 1.7μm BEHTM 45 Å, 1.8μm BEH 125 2.4μm 5.3μm 11.3μm Å Particles, an ACQUITYTM UPLCTM I-class PLUS System, either 1 × PBS (pH 7.4) or 100 mM phosphate buffer 8) phase. mode dominant when Weissenberg number (Wi=〈γ̇〉τR) much 10 average exceeds particle diameter. HDC, other hand, dominates Wi<1 (DNA chains remaining non-extended configuration). Maximum dsDNA resolution mixed HDC-SC approximately half work facilitates development methods characterizing various plasmid mixtures, containing linear, supercoiled, relaxed circular which all have different degree molecular stiffness.

Язык: Английский

Процитировано

9
Frit-Inlet Asymmetric Flow Field-Flow Fractionation for the Analysis of Lipid Nanoparticle-Protein Interactions DOI Creative Commons

Rand Abdulrahman,

Panida Punnabhum,

Robin Capomaccio

и другие.

Journal of Chromatography A, Год журнала: 2025, Номер 1743, С. 465663 - 465663

Опубликована: Янв. 11, 2025

Research into nanoparticle interactions with biomolecules has become increasingly important in nanomedicine. While lipid nanoparticles (LNPs) are widely used as drug delivery systems, there remains a gap understanding their fate circulation, which is crucial for selecting appropriate lipids during formulation development. This study the first to use Asymmetric Flow Field Fractionation (AF4) compare two types of LNPs: MC3-LNPs and SM-102-LNPs, model protein, bovine serum albumin (BSA). AF4 offers high-resolution separation, ability simultaneously perform multiparametric inline analysis multiple detectors. In this study, impact LNP size, morphology PDI on BSA corona formation were examined using multiangle light scattering (MALS) dynamic (DLS). separation revealed subpopulations MC3-LNPs, while SM102-LNPs exhibited single population. Analysis shape factor indicated 0.783 SM-102-BSA 0.741 0.795 (peak 1 2) MC3-BSA, confirming interaction between LNPs BSA. Both LNP-BSA induced aggregation. Overall, demonstrates effectiveness AF4, particularly when hyphenated multidetector separating from complex biological media studying LNP-protein interactions.

Язык: Английский

Процитировано

1
mRNA lipid nanoparticle formulation, characterization and evaluation DOI
Yutian Ma,

Rachel VanKeulen‐Miller,

Owen S. Fenton

и другие.

Nature Protocols, Год журнала: 2025, Номер unknown

Опубликована: Март 11, 2025

Язык: Английский

Процитировано

1
Rapid and Accurate Quantification of RNA in Lipid Nanoparticles by Scatter-Free UV/Visible Spectroscopy DOI
Eric C. Le Ru, Roland Böttger,

Darren T. Andrews

и другие.

Nano Letters, Год журнала: 2025, Номер unknown

Опубликована: Апрель 8, 2025

UV/visible spectroscopy is the method of choice for RNA quantification, thanks to its simplicity and accuracy. However, it cannot be used quantify in lipid nanoparticles (LNPs), such as those drug delivery mRNA vaccines, because light scattering by LNPs. Alternative methods RiboGreen fluorescence assay require much more sample preparation lack reproducibility Here we propose demonstrate an alternative approach using integrating sphere setup measure scatter-free absorption spectra. spectra RNA-loaded LNPs can then directly measured, total concentration deduced. The shows very good linearity precision (∼1.5%), accuracy estimated ∼5% when applied various mRNA-LNP formulations. This work paves way routine characterization payload LNP formulation research.

Язык: Английский

Процитировано

1
Analytical Ultracentrifugation to Assess the Quality of LNP-mRNA Therapeutics DOI Open Access
Giuditta Guerrini, Dóra Méhn, Diletta Scaccabarozzi

и другие.

International Journal of Molecular Sciences, Год журнала: 2024, Номер 25(11), С. 5718 - 5718

Опубликована: Май 24, 2024

The approval of safe and effective LNP-mRNA vaccines during the SARS-CoV-2 pandemic is catalyzing development next generation mRNA therapeutics. Proper characterization methods are crucial for assessing quality efficacy these complex formulations. Here, we show that analytical ultracentrifugation (AUC) can measure, simultaneously without any sample preparation step, sedimentation coefficients both formulation molecules. This allows measuring several attributes, such as particle size distribution, encapsulation efficiency density formulation. technique also be applied to study stability under stress conditions different buffers.

Язык: Английский

Процитировано

3
When conventional approach in toxicity assays falls short for nanomedicines: a case study with nanoemulsions DOI Creative Commons
Ines Nikolić, Jelena Đoković, Dóra Méhn

и другие.

Drug Delivery and Translational Research, Год журнала: 2025, Номер unknown

Опубликована: Янв. 8, 2025

Abstract The aim of this study was to assess the critical quality attributes parenteral nanoemulsion formulations by measuring several physicochemical parameters and linking them their in vitro performance, illustrating how simplistic routinely used approaches are insufficient for understanding a potential nanomedicine. Physicochemical characterization should encompass size distribution through at least two orthogonal techniques, such as dynamic light scattering (DLS) electron microscopy, with added value from analytical ultracentrifugation. In toxicity assessment performed using three different assays determine mitochondrial activity (WST-1), membrane integrity (lactate dehydrogenase release (LDH) assay), cell viability (propidium iodide (PI) staining). Special focus placed on estimating appropriate incubation times relevant results biological investigations. All had an average diameter around 100 nm. Conclusions regarding safety were assay-dependent: LDH PI-based showed good correlation, while WST-1 assay indicated that non-PEGylated formulation altered more significantly compared PEGylated ones. underlined selection cytotoxicity be based possible mechanism cellular perturbation. Alternatively, aspects tested. Additionally, there is need well-designed controls overcome nanoparticle effects avoid potentially false high results, which demonstrated. Combining orthogonal, standardized manner initial step reliable preclinical nanomedicines suggested. This represents key aspect new methodologies nanomedicine characterization. Graphical abstract

Язык: Английский

Процитировано

0
Therapeutic siRNA Quantification in Lipid Nanoparticles Using ion-pair reversed-phase Chromatography DOI Creative Commons
Ludivine Ferey,

Sandy Al Bardawil,

Emilie Dols

и другие.

Journal of Pharmaceutical and Biomedical Analysis Open, Год журнала: 2025, Номер unknown, С. 100072 - 100072

Опубликована: Март 1, 2025

Язык: Английский

Процитировано

0
Characterization of Lipid Nanoparticles Using Macro Mass Photometry: Insights into Size and Mass DOI Creative Commons
Jikang Wu, Yu Shrike Zhang, Zhijie Wu

и другие.

Analytica Chimica Acta, Год журнала: 2025, Номер unknown, С. 343944 - 343944

Опубликована: Март 1, 2025

Lipid nanoparticles (LNPs) have become an important delivery system for nucleic acids, as applied in the first RNAi drug and two COVID-19 mRNA vaccines approved by FDA. Despite advantages of their high cargo capacity, low immunogenicity allowing redosing, scalability low-cost manufacturing, challenges such liver accumulation difficulties quality control persist LNPs development. Conjugation antibodies or antibody fragments onto holds promise achieving precise targeting higher stability moieties on LNP surfaces. However, multiple-component products poses additional compared to alone. size mass are critical attributes which play roles determining LNPs' acid loading, conjugation level, biodistribution, capability, overall efficacy. Macro photometry (MMP), a single-molecule technique, enabled orthogonal characterization incorporating contrast analysis proxy particle combining it with measurement. This approach thus improves definition species within heterogeneous systems. Using MMP, this study revealed effects PEG-lipid concentration, encapsulation, mass. Additionally, demonstrated MMP's utility monitoring under various stress conditions, including pH, oxidation, freeze-thaw cycles, agitation. represents use analysis, simultaneously characterizing LNPs. It highlights potential MMP valuable tool supports formulation

Язык: Английский

Процитировано

0