Kinetics and Characterization of Non-enzymatic Fragmentation of Monoclonal Antibody Therapeutics

Pharmaceutical Research
Sahithi RavuluriAnurag S Rathore

Abstract

To understand non-enzymatic hydrolytic fragmentation of a monoclonal antibody therapeutic under temperature stressed conditions and investigating possible mechanism for the same. The mAb therapeutic was incubated at 50°C in phosphate buffer at pH 6.5 and fragmentation was monitored at different ionic strengths under stressed conditions. The incubated mAb was sampled at regular time intervals by analytical Size Exclusion Chromatography (SEC). It was observed that 57% of the mAb product fragmented over 4 days into two fragment species - Fc-Fab and Fab with molecular weights of 97 KDa and 47 KDa, respectively, as measured by mass spectrometry (MS) and sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The fragmentation rate was slow initially and then accelerated with time. No change in % aggregate level was observed in this duration, implying that degradation was primarily via fragmentation at high temperature. Kinetics of hydrolytic fragmentation was hypothesized and SEC data was fitted to estimate the kinetic rate constants. While degradation of the monomer into fragment species was non-Arrhenius with a negative activation energy, further degradation of Fab-Fc fragments into Fab or Fc fragments followed Arrh...Continue Reading

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Citations

Aug 30, 2019·Pharmaceutical Research·Rohit BansalAnurag S Rathore
Jun 20, 2021·Biomolecular Concepts·Rohit BansalNiraj Kumar Jha
Aug 31, 2021·European Journal of Pharmaceutics and Biopharmaceutics : Official Journal of Arbeitsgemeinschaft Für Pharmazeutische Verfahrenstechnik E.V·Shravan SreenivasanAnurag S Rathore

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Methods Mentioned

BETA
Size Exclusion Chromatography
Electrophoresis

Software Mentioned

Agilent Mass Hunter workstation
Thermo Scientific Chromeleon
Agilent Mass Hunter
Quantity One

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