(M1130-05-27) Evaluating the Efficacy of Polyoxyethylene Sorbitan as a Novel Surfactant in Biotherapeutic Antibody Formulations

Purpose: Our goal is to develop polyoxyethylene sorbitan as a novel surfactant to replace polysorbates for protein formulations with lipase activities. We generated polyoxyethylene sorbitan (POES, the hydrophilic part of polysorbates) and evaluated its effectiveness in protecting mAb formulations against agitation stress. We evaluated POES with two mAbs at low and high concentrations and compared it to polysorbates against agitation stress. The thermal stability of mAbs at long-term storage, accelerated and stress conditions in the presence of POES compared with polysorbate 80 (PS-80) were evaluated.

Methods: To prepare POES, we hydrolyzed polysorbates with 0.1M NaOH and neutralized with HCl. We isolated POES by removing free fatty acids by extraction with ethyl acetate. We performed agitation studies with mAb formulations with different concentrations of polysorbates or POES by either vortexing at 1000rpm for 2 hours or orbital shaking for 48 hours. We then measured protein aggregation (SEC), turbidity (OD₄₀₅) and protein recovery (RP-UPLC). We assessed thermal stability of mAbs in the presence of PS-80 or POES at long-term storage, accelerated and stress conditions (5, 25 and 40°C) for up to six months. We measured protein aggregation (SEC), charge variants (CEX), polysorbate concentration (CAD), pH, turbidity (OD₄₀₅), subvisible particle count (MFI) and protein recovery (RP-UPLC).

Results: The results of our agitation studies (Fig. 1) showed that for 2 h agitation at 1000 rpm both POES and PS-80 fully protected mAbs from agitation damage at 0.02-0.05%. Results for 48 h orbital shaking were similar and confirmed that POES protects the mAbs from aggregation with efficacy comparable to polysorbates. In addition, POES presents no risk of free fatty acid particle formation for mAb formulations with lipase activity as demonstrated by MFI data showing subvisible particle formation in the presence of POES significantly reduced compared to PS-80. Thermal stability data (Fig. 2) shows comparable or slightly lower soluble aggregate formation in mAb formulations in the presence of POES compared to PS-80. This is an indication that POES has no detrimental effect on thermal stability of mAb formulations.

Conclusion: POES has shown similar effectiveness to polysorbates in protecting mAb formulations from agitation stress. MAbs have also shown comparable thermal stability in the presence of POES and PS-80. Based on the data presented, POES can be further evaluated as a novel surfactant for antibody biotherapeutics with lipase activity.

Acknowledgements: CA, LB, FT are employees of Regeneron Pharmaceuticals.