A comparison of different orthogonal techniques for the investigation of the conformational stability of NIST mAb in liquid formulations

Sammanfattning: A great challenge during early development of medicinal antibodies is to determine the stability of the candidates. This is required to be able to select the candidates with the greatest developability. The stability of the candidates can be estimated using different orthogonal techniques and the aim of this investigation was thus to use orthogonal techniques for the investigation of conformational changes and aggregation of mAbs. The secondary aim was to obtain information concerning the effect that formulation components has on the stability of NIST mAb and which techniques that detects which conformational changes. The stability will be estimated for NIST mAb in liquid formulations where the mAb concentration, buffer type, tonicity, pH and storage time is altered. Differential scanning calorimetry (DSC), dynamic and static light scattering (DLS & SLS), circular dichroism (CD) and intrinsic fluorescence are the methods used to estimate the stability in the different formulations. During the investigations it was noted that the instruments detect different conformational changes as they investigate different parts of the mAb and they received different peak temperatures of the conformational changes. Most instruments detect conformational changes in the span of 54-78°C. The order of the instruments that detected the first conformational change, from the lowest to the highest, was: DLS, CD, intrinsic fluorescence and DSC. Additionally, the effect of the addition of salt was investigated using the nanoDSC and the Zetasizer and it was noticed that it caused destabilization of the mAb in both cases. The effect of pH on the mAb was investigated using DSC, DLS, CD and fluorescence. The CD measurements shows a positive linear trend between stability and pH for the investigated solutions, and a slight wavelength shift was detected during the fluorescence measurements between pH 5.2 and 5.7. Yet, the trend was not as clear for the DSC and it was unclear if DLS could detect a stability change based on the received results. Lastly, there are no clear correlations between the buffer types and the mAb stability for any of the investigated methods. Thus, it can be concluded that the methods detect different formulation changes, yet it is difficult to determine which investigational method is the most applicable in early formulation development