Developing Assays for Complex Biologics - Navigating the Challenges of the Next Generation of Therapeutics

The biologics landscape has undergone a fundamental transformation. Where assay development once centered on peptides and monoclonal antibodies — modalities with well-defined mechanisms of action and established analytical frameworks — today's pipeline looks markedly different. With over 3,500 gene, cell, and RNA therapies currently in preclinical or clinical development globally, encompassing in vivo CAR-T therapies, AAV-based gene therapies, antibody–RNA conjugates, and CRISPR-based modalities, the industry is confronting a level of mechanistic and biological complexity that existing bioassay frameworks were simply not designed to address. As these novel modalities advance toward regulatory submission, reliably measuring and assuring potency has emerged as one of the most pressing — and least resolved — challenges in the field. Unlike conventional biologics, complex modalities act through inherently multi-step mechanisms: requiring cellular uptake, intracellular trafficking, target engagement, and downstream functional activation before any measurable biological effect is observed. Multiple upstream steps must each succeed before a downstream signal is generated, and the cumulative efficiency of each step results in poor signal that undermines assay robustness. Unlike single-step mechanisms where the readout closely tracks the biological event of interest, complex multi-step pathways demand extensive design iteration — from cell system selection and assay format to signal amplification strategies — simply to achieve an assay that performs reliably. Using antibody–RNA conjugates (ARCs) as a working example, this talk examines the limitations of existing assay paradigms, the unique bioanalytical challenges imposed by next-generation modalities.