Process signatures in glatiramer acetate synthesis: structural and functional relationships

Abstract

Glatiramer Acetate (GA) is an immunomodulatory medicine approved for the treatment of multiple sclerosis, whose mechanisms of action are yet to be fully elucidated. GA is comprised of a complex mixture of polypeptides with different amino acid sequences and structures. The lack of sensible information about physicochemical characteristics of GA has contributed to its comprehensiveness complexity. Consequently, an unambiguous determination of distinctive attributes that define GA is of highest relevance towards dissecting its identity. Herein we conducted a study of characteristic GA heterogeneities throughout its manufacturing process (process signatures), revealing a strong impact of critical process parameters (CPPs) on the reactivity of amino acid precursors; reaction initiation and polymerization velocities; and peptide solubility, susceptibility to hydrolysis, and size-exclusion properties. Further, distinctive GA heterogeneities were correlated to defined immunological and toxicological profiles, revealing that GA possesses a unique repertoire of active constituents (epitopes) responsible of its immunological responses, whose modification lead to altered profiles. This novel approach established CPPs influence on intact GA peptide mixture, whose physicochemical identity cannot longer rely on reduced properties (based on complete or partial GA degradation), providing advanced knowledge on GA structural and functional relationships to ensure a consistent manufacturing of safe and effective products.

Sequencing Analysis and Identification of the Primary Peptide Component of the Dialyzable Leukocyte Extract “Transferon Oral”: The Starting Point to Understand Its Mechanism of Action

“…This suggests that in the cell lysis stage by freeze/thaw cycles, during the manufacturing process of Transferon, proteases from leukocytes are released into the medium and digest the exposed cellular components in a random but controlled pattern. This process resembles the stage of polymerization and acid digestion of glatiramer acetate peptides, where up to 10 36 peptides of an unknown sequence are generated, but with a consistent peptide polydispersion among batches ( Campos-Garcia et al, 2017 ).…”

An Immunomodulatory Peptide Dendrimer Inspired from Glatiramer Acetate

“…Glatiramer acetate (GA) is a random polypeptide of approximately 5–9 kDa composed of L‐alanine, L‐lysine, L‐glutamic acid and L‐tyrosine in a 4.2/3.4/1.4/1.0 ratio approximating the composition of myelin basic protein [1–5] . GA has been on the market since 1996 as one of the most successful first‐line treatments for multiple sclerosis (MS), a chronic autoimmune neurodegenerative disease [6–9] .…”

Accelerated Polypeptide Synthesis via N‐Carboxyanhydride Ring Opening Polymerization in Continuous Flow

“…An example of industrialized synthesis of polypeptides via NCA ROP is glatiramer acetate (tradename Copaxone for treatment of multiple sclerosis), obtained in batch mode. [13,14] Interestingly, the use of flow chemistry as a synthetic method is a strong emerging trend in the pharmaceutical industry; in fact, the Food and Drug Administration has been strongly stimulating the application of flow technology in the context of active pharmaceutical ingredients. [15] In response, the biopharmaceutical industry is beginning to recognize the advantages this technique can offer, and is adopting knowledge already present in academic labs, food, petrochemical, and chemical industries.…”