Research ArticlesImmunogenicity of aggregates of recombinant human growth hormone in mouse models
Section snippets
INTRODUCTION
Therapeutic proteins are susceptible to aggregation in response to a wide variety of stresses encountered during their manufacture, storage and delivery to patients.1 In turn, aggregates of therapeutic proteins may compromise their safety and efficacy.2, 3, 4, 5 The primary safety concern is that aggregates in therapeutic protein products may induce immune responses,6,7 which can have consequences ranging from reduction of product efficacy to patient fatality.8 In extreme cases, parenterally
Materials
The two commercial formulations of rhGH Nordiflex® (Novo Nordisk®, Bagsvaerd, Denmark) and Saizen® (Serono, Rockland, MA), were purchased from the University of Colorado apothecary, and are hereafter referred to as Product A and Product B, respectively. Sterile water for injection (SWFI) (Hospira, Inc., Lake Forest, IL) and 0.9% sodium chloride for injection (Hospira, Inc.) were also purchased form the University of Colorado apothecary. Histidine and mannitol were purchased from JT Baker
Stressing of rhGH Samples
The responses of the Product A and Product B formulations to the various stresses (agitation, freeze-thawing) were different. The agitated Product A samples were cloudy by the end of the 72 h of agitation whereas the agitated Product B samples were still clear. Similarly, the FT Product A samples began to become cloudy around the 12–15th freeze–thaw cycle; however, the FT Product B never showed signs of cloudiness even after the 20th freeze–thaw cycle. The contrast in aggregates produced in the
DISCUSSION
Parenteral administration of aggregates of a therapeutic protein can induce immune responses to the monomeric protein. However, little is known about the characteristics of aggregates that are capable of inducing immunogenicity and the mechanism by which they provoke the response.5 B cells can be stimulated to produce antibodies in T-cell independent mechanism that requires an antigen with a repetitive structure.39 Dintzis et al.40 determined that in order for polymeric antigens to activate
CONCLUSIONS
This study has shown that the immunogenicity of protein aggregates may depend on their size and the manner and solution conditions by which they are produced. Aggregates found in existing commercial formulations were immunogenic in the naïve adult and neonatally primed mice, as were aggregates generated by freeze-thawing- or agitation-induced stresses applied to the formulations, two types of stresses that are routinely encountered during production, handling, and storage of protein
Acknowledgements
Financial support for this study was provided by BaroFold, Inc., a company in which TWR and JFC hold financial interest.
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