7.2.3.4 Formulation changes

Biosimilarity: The FDA Perspective

274 may be found as traces in the finished product. Some impurities may be released by some compounds used for the capping process. These impurities may play the role of the amplifier for the immune response, even if they are not able to initiate an immune response themselves. Process-related impurities, including traces of residual DNA or proteins from the expression system, or contaminants that leach from the product container, can also influence the immunogenicity of recombinant biopharmaceuticals. Several examples exist in the modification of the recombinant protein’s immunogenicity with time, involving production of bacterial cells’ endotoxin level. DNA G-C patterns from bacteria or degraded proteins are able to activate toll-like receptors and act as adjuvants. (Toll-like receptors are a class of proteins playing a key role in the innate immune response. They are trans-membrane proteins containing receptors that detect danger signals located in the extracellular milieu, a trans-membrane medium, and an intracellular medium allowing the activation signal transduction.) However, the action of these impurities is limited to nonhuman proteins with a pseudovaccination activity. The adjuvants are unable to stimulate an immune response based on a T lymphocyte’s response independent of the tolerance breakdown of B lymphocytes.

7.2.3.4 Formulation changes Changes in formulations have been associated with ADAs. Two particularly interesting cases could illustrate how important the formulation and conservation conditions are. The first case concerns EPO. Cases of PRCA after treatment with EPO are known, but rare. Anti-EPO antibodies are formed exogenously as well as endogenously, after administration of recombinant human EPO (rHuEPO). These cases have occurred after a changed in the formulation of the finished product, with human albumin used as a stabilizer replaced by polysorbate 80. Different hypotheses to explain the immune system tolerance breakdown after administration of rHuEPO lead, among other things, to the impurities’ extraction from the syringe’s plunger rod stopper, playing booster in the presence of EPO. During the analysis of batches called into question, no increase in the level of aggregates or in the level of truncated or degraded EPO has been evidenced. In that case, there must have been several factors having fostered the immune system’s tolerance breakdown. In particular, the subcutaneous route of administration may be incriminated, as it is known to produce a pseudovaccination effect. The second case involves the conservation conditions of a freeze-dried formulation of interferon a-2a (rHuINF a-2a) that has been stabilized by human albumin. At room temperature, a partial oxidation of rHuINF a-2a has made the formation of aggregates easier with intact interferon and albumin. These aggregates led to the therapeutic preparation’s immunogenicity. These cases illustrate how important the finished product formulation study and the evaluation that has to be done of the possible consequences