Performance Comparison of Spectral Distance Calculation Methods

Circular dichroism (CD) spectroscopy is a widely used technique for assessing the higher-order structure (HOS) of biopharmaceuticals, including antibody drugs. Since the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use established quality control guidelines, objective evaluation of spectral similarity has been required in order to assess structural comparability. Several spectral distance quantification methods and weighting functions to increase sensitivity have been proposed, but not many reports have compared their performance for CD spectra. We constructed comparison sets that combine actual spectra and simulated noise and performed a comprehensive performance evaluation of each spectral distance calculation method and weighting function under conditions that consider spectral noise and fluctuations from pipetting errors. The results showed that using the Euclidean distance or Manhattan distance with Savitzky-Golay noise reduction is effective for spectral distance assessment. For the weighting function, it is preferable to combine the spectral intensity weighting function and the noise weighting function. In addition, the introduction of the external stimulus weighting function should be considered to improve the sensitivity. It is crucial to select the weighting function based on the balance between spectral changes and noise distributions for robust, sensitive antibody HOS similarity assessment.