PXD011017

PXD011017 is an original dataset announced via ProteomeXchange.

Title	Dilute-and-Shoot Analysis of Therapeutic Monoclonal Antibody Variants in Fermentation Broth: A Method Capability Study
Description	Monoclonal antibodies (mAbs) are widely applied as highly specific and efficient thera-peutic agents for various diseases, including cancer, inflammatory and autoimmune diseases. As protein production in cellular systems inherently generates a multitude of molecular variants, manufacturing of mAbs requires stringent control in order to ensure safety and efficacy of the drug. Moreover, monitoring of mAb variants in the course of the fermentation process may allow instant tuning of process parameters to maintain optimal cell culture conditions. Here, we describe a fast and robust workflow for the characterization of mAb variants in fermentation broth. Sample preparation is minimal in that the fermentation broth is shortly centrifuged before dilution and HPLC-MS analysis in a short 15-min gradient run. In a single analysis, N-glycosylation and truncation variants of the expressed mAb can be identified at the intact protein level. The molecular attributes of the expressed therapeutic protein may thus be continuously monitored to ensure the desired product profile. Simultaneously, absolute quantification of mAb content in fermentation broth yielded concentrations of 67 and 2578 ng.mL-1 at the beginning and end of fermentation, respectively. The whole workflow features excellent robustness as well as retention time and peak area stability of 0.3% and 4.9% RSD. Additional enzymatic removal of N-glycans enables determination of mAb glycation levels, which are subsequently considered in relative N-glycoform quantification to correct for isobaric galactosylation. Application of the described workflow in an industrial environment may therefore substantially enhance in-process control in mAb production as well as targeted biosimilar development.
HostingRepository	PRIDE
AnnounceDate	2019-11-12
AnnouncementXML	Submission_2019-11-12_13:49:38.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Christof Regl
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	complex glycosylation
Instrument	Q Exactive

Revision	Datetime	Status	ChangeLog Entry
0	2018-09-05 23:59:44	ID requested
⏵ 1	2019-11-12 13:49:40	announced

Regl C, Wohlschlager T, Esser-Skala W, Wagner I, Samonig M, Holzmann J, Huber CG, Dilute-and-shoot analysis of therapeutic monoclonal antibody variants in fermentation broth: a method capability study. MAbs, 11(3):569-582(2019) [pubmed]

ProteomeXchange project tag: Antibodies (B/D-HPP), Biology/Disease-Driven Human Proteome Project (B/D-HPP), Human Proteome Project

curator keyword: Technical

submitter keyword: multiple attribute monitoring, MAM, monoclonal antibodies, high-performance liquid chromatography, electrospray ionization mass spectrometry, fermentation, antibody subunits, glycation, glycosylation, structural analysis, biosimilar, process analytical technology, PAT, process monitoring

Christian G. Huber
contact affiliation	1: Department of Molecular Biology, Division of Chemistry andBioanalytics, University of Salzburg, Hellbrunner Strasse 34, 5020 Salzburg, Austria 2: Christian Doppler Laboratory for Innovative Tools for Biosimilar Characterization, University of Salzburg, Hellbrunner Strasse 34, 5020 Salzburg, Austria
contact email	c.huber@sbg.ac.at
lab head
Christof Regl
contact affiliation	University of Salzburg
contact email	christof.regl@sbg.ac.at
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD011017
     1. Label: PRIDE project
     2. Name: Dilute-and-Shoot Analysis of Therapeutic Monoclonal Antibody Variants in Fermentation Broth: A Method Capability Study