Developability studies before initiation of process development: improving manufacturability of monoclonal antibodies

Abstract

Monoclonal antibodies constitute a robust class of therapeutic proteins. Their stability, resistance to stress conditions and high solubility have allowed the successful development and commercialization of over 40 antibody-based drugs. Although mAbs enjoy a relatively high probability of success compared with other therapeutic proteins, examples of projects that are suspended due to the instability of the molecule are not uncommon. Developability assessment studies have therefore been devised to identify early during process development problems associated with stability, solubility that is insufficient to meet expected dosing or sensitivity to stress. This set of experiments includes short-term stability studies at 2-8 þC, 25 þC and 40 þC, freeze-thaw studies, limited forced degradation studies and determination of the viscosity of high concentration samples. We present here three case studies reflecting three typical outcomes: (1) no major or unexpected degradation is found and the study results are used to inform early identification of degradation pathways and potential critical quality attributes within the Quality by Design framework defined by US Food and Drug Administration guidance documents; (2) identification of specific degradation pathway(s) that do not affect potency of the molecule, with subsequent definition of proper process control and formulation strategies; and (3) identification of degradation that affects potency, resulting in program termination and reallocation of resources.

Keywords: CQA; QbD; analytics; developability; development; discovery; forced degradation; process control; stability.

Figure 1. Overlay of IEX-HPLC chromatograms of mAb A samples stored for one month (A) and three months (B) at -80°C, 2–8°C, 25°C and 40°C. Overlay of IEX-HPLC chromatograms of mAb A samples challenged with high and low pHs for 48 h at 40°C (C) and light induced (0.2xICH) stress (D).

Figure 2. Overlay of IEX-HPLC chromatograms of mAb B samples stored for one month (A) and three months (B) at -80°C, 2–8°C, 25°C and 40°C. Evolution during stability studies of relative amount of deamidation at HCAsn55 (N55, blue trace) and mAb B potency (red trace) (C). Overlay of IEX-HPLC chromatograms of mAb B samples stored for one month at 40°C at pH 4.5 and pH 6.0 (D). Thermal unfolding profile of high concentration solution (190 mg/ml) of mAb B (E).

Figure 3. Overlay of IEX-HPLC chromatograms of mAb C samples stored for one month (A) and three months (B) at -80°C, 2–8°C, 25°C and 40°C. Evolution during stability studies of relative amount of deamidation at HCAsn33 (N33, blue trace) and mAb B potency (red trace) (C). The bar graph represents the relative area of acidic variants after one month at 40°C in six different formulations (D). Deamidation at Asn33 in rhesus monkey serum stability studies (E): TIC of the doubly charged ion of peptide 25–39 at day 0, 7, 14 and 21: The peak at ~33–34 min is the early eluting non deamidated parent peptide (Asn33), the two peaks eluting at 34–36 and 35–37 min are the deamidated product peptides (iso-Asp33 and Asp33, respectively). At all-time points the ratio of deamidated product peptides IsoAsp/Asp is the same, ~1:5 throughout the whole experiment. Titration of mAb C in rhesus monkeys serum by an anti-kappa light chain (PK curve) and antigen ligand-capture (Antigen curve) ELISAs after second injection (F).