The Use of Mass Spectrometry in Therapeutic Protein Biologics License Applications: A Retrospective Review Revisited.

Abstract

Biologic license applications (BLAs) for 93 therapeutic proteins approved between 2016 and 2020 were analyzed for use of mass spectrometry (MS) as a follow up to a previous study that assessed MS use in BLAs from 2000 to 2015. Thirty percent of these BLAs were biosimilars, while only one biosimilar BLA was approved prior to 2016. This analysis evaluated the use of a variety of MS techniques and instrumentation. Results were further interpreted based on the relationship of MS use over time, between drug types, and between new drugs and biosimilars. MS data were included in 93 BLAs examined. The top eight quality attributes most assessed by MS in rank order were amino acid sequence, molecular mass, oxidation, disulfide bonds, deamidation, glycosylation, N-terminal sequence variants, and C-terminal sequence variants. These attributes were the same top attributes seen previously from BLAs approved between 2000 and 2015, and the use of MS to analyze them generally continued to increase across the new time frame. The average number of attributes analyzed by MS per BLA also continued to increase over the extended time frame of 21 years. High-resolution, accurate mass instrumentation such as the Orbitrap and time-of-flight (TOF) usage increased over time for all assessed attributes, while matrix-assisted laser desorption/ionization (MALDI)-TOF/(TOF) usage decreased. From highest to lowest rank, the top 11 attributes were antibody drug conjugate (ADC) characterization (i.e., drug load distribution/drug to antibody ratio (DAR), ADC and linkage site, and synthetic linker), isomerization, folding/higher-order structure (HOS), truncation, host cell proteins (HCPs), sequence variants (amino acid substitutions), succinimidation, glycation, PEGylation, charge variants, and oxidation.