A Precise qNMR Method for the Rapid Quantification of Lot-to-Lot Variations in Multiple Quality Attributes of Pentosan Polysulfate Sodium

Wang, Kai; Dai, Weixiang; Qian, Keduo; Scott, Barbara; Chen, Kang

doi:10.1208/s12248-023-00815-4

AAPS J

2023

DOI: 10.1208/s12248-023-00815-4

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A Precise qNMR Method for the Rapid Quantification of Lot-to-Lot Variations in Multiple Quality Attributes of Pentosan Polysulfate Sodium

Kai Wang

Weixiang Dai

Keduo Qian

et al.

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2024

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Cited by 2 publications

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An Accurate and Fast ³¹P qNMR Assay Method for Oligonucleotide Therapeutics

Li,

Chen,

Zhang

et al. 2024

Anal. Chem.

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Chemically modified nucleic acid molecules have been developed as oligonucleotide therapeutics, and its assay is critical in quality assurance. The common DNA/RNA quantification method using UV-260 nm can lack accuracy because of structure modifications and the possible formation of higher-order structure (HOS). Additionally, process-associated water and counterions affect the accuracy in gravimetric analysis. Thus, to improve accuracy, efficiency, and flexibility, in this work a fast (<1 h) externally referenced 31 P quantitative-NMR (qNMR) method was developed. The qNMR assay results agreed within 1−5% of the UV-260 nm results for the single-stranded DNA standards, confirming the method accuracy. Next, an NMR and UV comparison study was performed on intact oligonucleotide drug products. The 31 P qNMR method showed 7 ± 2%, 8 ± 1%, and 12 ± 1% lower concentration values compared with drug product labels for eteplirsen, inotersen, and inclisiran, respectively. Meanwhile the UV-260 nm results showed 28 ± 3%, 10 ± 3%, and 10 ± 1% lower concentrations than the label for the same three drugs. The agreement between NMR and UV for phosphorothioate (PS)-based inotersen and mostly phosphodiester (PO)-based inclisiran suggest that the labeled concentration may have been obtained using different extinction coefficients. The underestimate of UV results for eteplirsen, which has a phosphorodiamidate morpholino oligomer (PMO) structure, suggests that the UV-260 nm extinction coefficient may need to be re-established for the PMO based oligonucleotide. Therefore, the 31 P qNMR method could be a primary assay method for the oligonucleotide drug and reference standard.

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An Accurate and Fast ³¹P qNMR Assay Method for Oligonucleotide Therapeutics

Li,

Chen,

Zhang

et al. 2024

Anal. Chem.

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Intact NMR Approach Quickly Reveals Synchronized Microstructural Changes in Oil-in-Water Nanoemulsion Formulations

Wang,

Li,

Chen

2024

AAPS J

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A soft-core oil-in-water (o/w) nanoemulsion (NE) is composed of nanometer (nm) sized oil droplets, stabilized by a surfactant layer and dispersed in a continuous bulky water phase. Characterization of the o/w NE molecule arrangements non-invasively, particularly the drug phase distribution (DPD) and its correlation to oil globule size (OGS), remains a challenge. Here we demonstrated the analytical methods of intact 19F Nuclear Magnetic Resonance (NMR) and 1H diffusion ordered spectroscopy (DOSY) NMR for their specificity in measuring DPD and OGS, respectively, on three NE formulations containing the active ingredient difluprednate (DFPN) at the same concentration. The results illustrated synchronized molecular rearrangement reflected in the DPD and OGS upon alterations in formulation. Addition of surfactant resulted in a higher DPD in the surfactant layer, and concomitantly smaller OGS. Mechanic perturbation converted most of the NE globules to the smaller thermodynamically stable microemulsion (ME) globules, changing both DPD and OGS to ME phase. These microstructure changes were not observed using 1D 1H NMR; and dynamic light scattering (DLS) was only sensitive to OGS of ME globule in mechanically perturbed formulation. Collectively, the study illustrated the specificity and essential role of intact NMR methods in measuring the critical microstructure attributes of soft-core NE systems quickly, accurately, and non-invasively. Therefore, the selected NMR approach can be a unique diagnostic tool of molecular microstructure or Q3 property in o/w NE formulation development, and quality assurance after manufacture process or excipient component changes. Graphical abstract

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A Precise qNMR Method for the Rapid Quantification of Lot-to-Lot Variations in Multiple Quality Attributes of Pentosan Polysulfate Sodium

Cited by 2 publications

References 25 publications

An Accurate and Fast ³¹P qNMR Assay Method for Oligonucleotide Therapeutics

An Accurate and Fast ³¹P qNMR Assay Method for Oligonucleotide Therapeutics

Intact NMR Approach Quickly Reveals Synchronized Microstructural Changes in Oil-in-Water Nanoemulsion Formulations

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A Precise qNMR Method for the Rapid Quantification of Lot-to-Lot Variations in Multiple Quality Attributes of Pentosan Polysulfate Sodium

Cited by 2 publications

References 25 publications

An Accurate and Fast 31P qNMR Assay Method for Oligonucleotide Therapeutics

An Accurate and Fast 31P qNMR Assay Method for Oligonucleotide Therapeutics

Intact NMR Approach Quickly Reveals Synchronized Microstructural Changes in Oil-in-Water Nanoemulsion Formulations

Contact Info

Product

Resources

About

An Accurate and Fast ³¹P qNMR Assay Method for Oligonucleotide Therapeutics

An Accurate and Fast ³¹P qNMR Assay Method for Oligonucleotide Therapeutics