(480f) The Design and Synthesis of Novel Excipients for the Suppression of Protein Aggregation

Due to the advancement of recombinant DNA technology and cell biology, the past two decades have seen an explosive growth in protein based therapeutics, e.g. monoclonal antibodies. Over 130 protein based therapeutics have been approved by the FDA for clinical use and well over 200 more are in some phase of development. However, rapid commercialization of these products has not been fully realized due to the presence of many barriers, namely the physical and chemical instabilities of proteins. Of these degradation pathways, protein aggregation is arguably the most common and troubling manifestation of protein instability, occurring in almost all phases of development. In addition to reducing efficacy, if administered to a patient, aggregates can cause adverse reactions, such as immune response, sensitization, or even anaphylactic shock. The structural differences among different proteins are so significant that the application of a universal stabilization strategy has not yet been successful, though the effects of common excipients are generally universal (e.g. sugars increase the free energy of unfolding thus deter the formation of aggregation prone species). The current approach to stabilizing protein drugs against aggregation is by trial-and-error testing of different combinations of cosolutes (e.g. salts, sugars, surfactants, arginine, etc.) using empirically derived heuristics. While ubiquitously used, this approach is inefficient and does not always enable the discovery of stable protein solution formulations. Thus many products must be lyophilized and reconstituted prior to injection, which is highly undesirable.

In response to this major problem, we have developed a theory for how a novel class of excipients, termed ?neutral crowders?, should inhibit protein association reactions. This new class of excipients has the potential for wide spread application as a universal stabilizer of biopharmaceuticals. The basis for the mechanism of a ?neutral crowder? is that a large additive which is neither excluded from nor bound to the surface of the protein will solvate the protein much like water but will tend to deter association due to the unfavorable entropic effect arising when the large additive is forced to be excluded from the gap formed between two associating protein molecules. To explore the possibility of such additives, we have synthesized a wide variety of large compounds with different surface groups known to bind to the surface of the protein in the hopes of developing an additive that has the right balance of attraction and steric exclusion, and thus behave as a ?neutral crowder?. So far we have had some initial success at developing such excipients and at inhibiting aggregation, allowing us to refine our designs for developing excipients that will significantly slow aggregation.