title: The use of generic surrogate peptides for the quantitative analysis of human immunoglobulin G1 in pre-clinical species with high-resolution mass spectrometry.
abstract: In the present study, the application of a liquid chromatography high-resolution mass spectrometry (LC-HRMS) analytical assay for the quantitative analysis of a recombinant human immunoglobulin G1 (hIgG1) in rat serum is reported using three generic peptides GPSVFPLAPSSK (GPS), TTPPVLDSDGSFFLYSK (TTP), and VVSVLTVLHQDWLNGK (VVS). Moreover, the deamidation site of a fourth peptide FNWYVDGVEVHNAK (FNW) was identified and further excluded from the assay evaluation due to the inaccuracy of the quantitative results. The rat serum samples were spiked with a fully labeled hIgG1 as internal standard (ISTD). The digestion with trypsin was performed onto the pellet prior to peptide analysis by LC-HRMS using a quadrupole time of flight (QTOF) mass analyzer operating in selected reaction monitoring (SRM) mode with enhanced duty cycles (EDC). The assay linearity for the three investigated peptides was established for a hIgG1 (hIgG1A) from 1.00 to 1000 μg mL(-1) with a mean coefficient of determination (R (2)) higher than 0.9868. The inter-day accuracy and precision obtained in rat serum over 3 days were ≤11.4 and ≤10.5%, respectively. Short-term stability on the auto-sampler at 6 °C for 30 h, at RT for 48 h, and a 100-fold dilution factor were demonstrated. In addition, QC samples prepared in cynomolgus monkey serum and measured with the present method met the acceptance criteria of ±20.0 and ≤20.0% for all three peptides regarding accuracy and precision, respectively. The LC-HRMS method was applied to the analysis of samples from five individual cynomolgus monkeys dosed with a second hIgG1 (hIgG1B) and consistent data were obtained compared to the LC-MS/MS method (conventional triple quadrupole (QqQ) mass analyzer operating in SRM). The present data demonstrate that LC-HRMS can be used for the quantitative analysis of hIgG1 in both species and that quantification is not only limited to classical QqQ instruments.
authors.affiliation: Drug Metabolism and Pharmacokinetics, Novartis Institutes for Biomedical Research, Fabrikstrasse 14 - Novartis Campus, 4056, Basel, Switzerland.|Laboratoire de Spectrométrie de Masse BioOrganique, Institut Pluridisciplinaire Hubert Curien, CNRS-Université de Strasbourg, UMR 7178, 25 rue Becquerel, 67087, Strasbourg, France.
authors.forename: Christian
authors.initials: C
authors.lastname: Lanshoeft
authors.affiliation: Drug Metabolism and Pharmacokinetics, Novartis Institutes for Biomedical Research, Fabrikstrasse 14 - Novartis Campus, 4056, Basel, Switzerland.
authors.forename: Thierry
authors.initials: T
authors.lastname: Wolf
authors.affiliation: Drug Metabolism and Pharmacokinetics, Novartis Institutes for Biomedical Research, Fabrikstrasse 14 - Novartis Campus, 4056, Basel, Switzerland. olivier.heudi@novartis.com.
authors.forename: Olivier
authors.initials: O
authors.lastname: Heudi
authors.affiliation: Laboratoire de Spectrométrie de Masse BioOrganique, Institut Pluridisciplinaire Hubert Curien, CNRS-Université de Strasbourg, UMR 7178, 25 rue Becquerel, 67087, Strasbourg, France.
authors.forename: Sarah
authors.initials: S
authors.lastname: Cianférani
authors.affiliation: Drug Metabolism and Pharmacokinetics, Novartis Institutes for Biomedical Research, Fabrikstrasse 14 - Novartis Campus, 4056, Basel, Switzerland.
authors.forename: Samuel
authors.initials: S
authors.lastname: Barteau
authors.affiliation: Drug Metabolism and Pharmacokinetics, Novartis Institutes for Biomedical Research, Fabrikstrasse 14 - Novartis Campus, 4056, Basel, Switzerland.
authors.forename: Markus
authors.initials: M
authors.lastname: Walles
authors.affiliation: Drug Metabolism and Pharmacokinetics, Novartis Institutes for Biomedical Research, Fabrikstrasse 14 - Novartis Campus, 4056, Basel, Switzerland.
authors.forename: Franck
authors.initials: F
authors.lastname: Picard
authors.affiliation: Drug Metabolism and Pharmacokinetics, Novartis Institutes for Biomedical Research, Fabrikstrasse 14 - Novartis Campus, 4056, Basel, Switzerland.
authors.forename: Olivier
authors.initials: O
authors.lastname: Kretz
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods / Q000592:standards; D005260:Female; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood* / Q000235:genetics; D008252:Macaca fascicularis; D013058:Mass Spectrometry / Q000379:methods* / Q000592:standards; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000097:blood* / Q000235:genetics / Q000494:pharmacology; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods
chemical_list: D007074:Immunoglobulin G; D010455:Peptides; D011994:Recombinant Proteins
keywords: Deamidation; Generic peptide; High-resolution mass spectrometry; Immunoglobulin G; Pellet digestion; Pre-clinical species