Biophysical and Structural Characterization of Antibody-Drug Conjugates.

Area of Science:

• Biochemistry
• Biotechnology
• Pharmaceutical Sciences

Background:

• Antibody-drug conjugates (ADCs) are targeted therapeutics combining monoclonal antibodies with cytotoxic payloads.
• ADCs offer specificity but face challenges like aggregation, misfolding, and immunogenicity, impacting efficacy and pharmacokinetics.
• Early assessment of ADC developability is crucial to mitigate risks before extensive studies.

Purpose of the Study:

• To review established biophysical and structural techniques for ADC characterization.
• To introduce emerging methods for assessing ADC developability and potential liabilities.
• To guide efficient development of ADCs and next-generation protein therapeutics.

Main Methods:

• Stability assessment using Differential Scanning Calorimetry (DSC), Differential Scanning Fluorimetry (DSF), and spectroscopy (CD, intrinsic fluorescence).
• Drug-to-antibody ratio (DAR) measurement via absorbance spectroscopy and chromatography.
• Aggregation analysis using Dynamic Light Scattering (DLS), Static Light Scattering (SLS), SEC-MALS, FCS, and Analytical Ultracentrifugation (AUC).
• Mass spectrometry (MS) and Hydrogen-Deuterium Exchange MS (HDX-MS) for stability, modifications, and dynamics.

Main Results:

• Various biophysical techniques effectively characterize critical ADC attributes like stability, DAR, and aggregation.
• Mass spectrometry provides deep insights into ADC structure, modifications, and dynamics.
• Emerging techniques and AI/ML show promise for accelerated ADC development.

Conclusions:

• Biophysical and structural characterization are essential for understanding ADC developability and liabilities.
• A combination of established and novel techniques enables comprehensive ADC assessment.
• Future advancements in biophysical assays and AI/ML will expedite the development of ADCs and protein therapeutics.