Concerted Application of LC–MS and Ligand Binding Assays to Better Understand Exposure of a Large Molecule Drug

Xu, Weifeng; Jiang, Hao; Titsch, Craig; Gadkari, Snaehal; Batog, Alicja; Wang, Bonnie; Hippeli, Lauren; Yamamoto, Brent; Chadwick, Kristina D.; Wheeler, Jennifer; Thompson, Chris; Stahl, James E.; Willett, Scott; DeSilva, Binodh; Myler, Heather; Dodge, Robert; Pillutla, Renuka

doi:10.4155/bio-2018-0108

Cited by 3 publications

(6 citation statements)

References 14 publications

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“…Traditionally, ligand-binding assays (LBAs) and triple quadrupole-based assays are employed for the quantitative detection of large molecules [38]. However, the above two methods have their shortcomings in the quantification of macromolecules, which can be solved by liquid chromatography (LC)-HRMS.…”

Section: Discussionmentioning

confidence: 99%

Quantitative Measurement of Melittin in Asian Honeybee Venom Using a New Method Including UPLC-QqTOF-MS

Huang

Wang

Guo

et al. 2020

Toxins

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Asian honeybee venom is widely used in traditional oriental medicine. Melittin is the main component of Asian honeybee venom. In the present study, an ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QqTOF-MS) method was used for accurate qualitative and quantitative analyses of melittin in Asian honeybee venom. The results showed that the dynamic linear range of melittin was from 0.094 to 20 μg/mL, and the limit of quantification was 0.3125 μg/mL. The spiking recovery of melittin in honeybee venom ranged from 84.88% to 93.05%. Eighteen Asian honeybee venom samples in eighteen batches were collected from two different zones of China, and their melittin contents were measured. The contents of melittin in Asian honeybee venom samples was 33.9–46.23% of dry weight. This method proved a useful tool for the rapid evaluation of the authenticity and quality of Asian honeybee venom in terms of the melittin contents, and will contribute to a broader understanding of Asian honeybee venom.

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Section: Discussionmentioning

confidence: 99%

Quantitative Measurement of Melittin in Asian Honeybee Venom Using a New Method Including UPLC-QqTOF-MS

Huang

Wang

Guo

et al. 2020

Toxins

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“…Distinguishing zinpentraxin alfa from the preexisting SAP in circulation is key to accurate exposure assessment of the therapeutic protein during the IPF treatment in clinical studies. Previously reported ELISA and LC-MS methods were applied to quantification of zinpentraxin alfa in nonclinical studies where no interference from human endogenous SAP was expected (17,18). However, neither assay was amenable to differentiate zinpentraxin alfa from SAP in clinical studies, i.e., the LC-MS was based on a common surrogate tryptic peptide shared by both isomers (17) and the ELISA utilized reagents lacking selective structural recognition capability (18), respectively.…”

Section: Discussionmentioning

confidence: 99%

“…An immunoaffinity liquid chromatography mass spectrometry (LC-MS) method and an enzyme-linked immunosorbent assay (ELISA) were reported for quantification of SAP and zinpentraxin alfa. Immunoaffinity LC-MS based on a selected zinpentraxin alfa signature peptide was reported to measure circulating zinpentraxin alfa in rat and monkey plasma samples, which did not contain endogenous human SAP (17). The surrogate peptide selected for quantification is shared by both zinpentraxin alfa and SAP (17).…”

Section: Introductionmentioning

confidence: 99%

“…Immunoaffinity LC-MS based on a selected zinpentraxin alfa signature peptide was reported to measure circulating zinpentraxin alfa in rat and monkey plasma samples, which did not contain endogenous human SAP (17). The surrogate peptide selected for quantification is shared by both zinpentraxin alfa and SAP (17). Therefore, this non-specific LC-MS approach is not applicable to measure zinpentraxin alfa in human samples.…”

Section: Introductionmentioning

confidence: 99%

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Novel Selective Quantification of Zinpentraxin Alfa Biotherapeutic in the Presence of Endogenous Isomer in Plasma Samples of Idiopathic Pulmonary Fibrosis Patients Using Immunoaffinity LC–MS

Li,

Arjomandi,

Sun

et al. 2023

AAPS J

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Idiopathic pulmonary fibrosis (IPF) is a progressive fatal interstitial lung disease that affects three million patients worldwide and currently without an effective cure. Zinpentraxin alfa, a recombinant human pentraxin-2 (rhPTX-2) protein, has been evaluated as a potential drug candidate for the treatment of IPF. Clinical pharmacokinetic analysis of zinpentraxin alfa has been challenging historically due to interference from serum amyloid P component (SAP), an endogenous human pentraxin-2 protein. These molecules share an identical primary amino acid sequence and glycan composition; however, zinpentraxin alfa possesses α2,3-linked terminal sialic acid residues while SAP is an α2,6-linked isomer. By taking advantage of this only structural difference, we developed a novel assay strategy where α2,3-sialidase was used to selectively hydrolyze α2,3-linked sialic acid residues, resulting in desialylated zinpentraxin alfa versus unchanged sialylated SAP, following an immunoaffinity capture step. Subsequent tryptic digestion produced a unique surrogate asialo-glycopeptide from zinpentraxin alfa and allowed specific quantification of the biotherapeutic in human plasma. In addition, a common peptide shared by both molecules was selected as a surrogate to determine total hPTX-2 concentrations, i.e., sum of zinpentraxin alfa and SAP. The quantification methods for both zinpentraxin alfa and total hPTX-2 were validated and used in pharmacokinetic assessment in IPF patients. The preliminary results suggest that endogenous SAP levels remained largely constant in IPF patients throughout the treatment with zinpentraxin alfa. Our novel approach provides a general bioanalytical strategy to selectively quantify α2,3-sialylated glycoproteins in the presence of their corresponding α2,6-linked isomers. Graphical Abstract

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“…55 LC-MS is gaining popularity for being highly selective and possessing multiplexing ability, whereas LBA performs better in terms of throughput and sensitivity. [56][57][58][59] A decision-tree that helps to select the right assay and three case studies using this approach were presented. Some factors to considers are: limit of quantification to be achieved, moieties to be quantified (total drug vs free drug) and whether in vivo integrity of the compound must be confirmed.…”

Section: Session 6: Non-canonical Biologics Formats: Challenges In Bioanalytics Pkpd and Biotransformation For Complex Biologics Formatsmentioning

confidence: 99%

Challenges of non-clinical safety testing for biologics: A Report of the 9th BioSafe European Annual General Membership Meeting

Kissner

Blaich

Baumann

et al. 2021

mAbs

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New challenges and other topics in non-clinical safety testing of biotherapeutics were presented and discussed at the nineth European BioSafe Annual General Membership meeting in November 2019. The session topics were selected by European BioSafe organization committee members based on recent company achievements, agency interactions and new data obtained in the non-clinical safety testing of biotherapeutics, for which data sharing would be of interest and considered as valuable information. The presented session topics ranged from strategies of in vitro testing, immunogenicity prediction, bioimaging, and developmental and reproductive toxicology (DART) assessments to first-in-human (FIH) dose prediction and bioanalytical challenges, reflecting the entire space of different areas of expertise and different molecular modalities. During the 9 th meeting of the European BioSafe members, the following topics were presented and discussed in 6 main sessions (with 3 or 4 presentations per session) and in three small group breakout sessions: 1) DART assessment with biotherapeutics: what did we learn and where to go?; 2) Non-animal testing strategies; 3) Seeing is believing: new frontiers in imaging; 4) Predicting immunogenicity during early drug development: hope or despair?; 5) Challenges in FIH dose projections; and 6) Non-canonical biologics formats: challenges in bioanalytics, PKPD and biotransformation for complex biologics formats. Small group breakout sessions were organized for team discussion about 3 specific topics: 1) Testing of cellular immune function in vitro and in vivo; 2) MABEL approach (toxicology and pharmacokinetic perspective); and 3) mRNA treatments. This workshop report presents the sessions and discussions at the meeting.

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Concerted Application of LC–MS and Ligand Binding Assays to Better Understand Exposure of a Large Molecule Drug

Abstract: The resultant LC-MS data confirmed the original data and the lack of dose-dependent exposure is now understood to be due to the multiple and diverse targets and functions and resultant complex biodistribution rather than an assay artifact.

Cited by 3 publications

References 14 publications

Quantitative Measurement of Melittin in Asian Honeybee Venom Using a New Method Including UPLC-QqTOF-MS

Quantitative Measurement of Melittin in Asian Honeybee Venom Using a New Method Including UPLC-QqTOF-MS

Novel Selective Quantification of Zinpentraxin Alfa Biotherapeutic in the Presence of Endogenous Isomer in Plasma Samples of Idiopathic Pulmonary Fibrosis Patients Using Immunoaffinity LC–MS

Challenges of non-clinical safety testing for biologics: A Report of the 9th BioSafe European Annual General Membership Meeting

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