High-throughput optimization of antibody production in CHO cells by tuning heavy- and light-chain promoter strength

TL;DR

This paper introduces a high-throughput method to optimize antibody production in CHO cells by testing different promoter strengths for heavy and light chains.

Contribution

A scalable, transient screening platform using design-of-experiments to identify optimal LC/HC promoter ratios for antibody production.

Findings

Optimal LC/HC promoter ratios are antibody-specific and can significantly affect titer and cell viability.

High LC with medium HC expression often maximizes titer while maintaining cell health for some antibodies.

Low-strength promoters for either chain consistently result in poor antibody production.

Abstract

Monoclonal antibody (mAb) production in CHO cells depends, among other factors, on balanced co-expression of the heavy (HC) and light (LC) chains. Imbalances between HC and LC can reduce titer, compromise product quality, and negatively affect cell viability, which means that the optimal LC/HC expression ratio should be identified as early as possible in cell line development. However, systematically testing multiple LC/HC expression ratios for many antibody candidates using traditional workflows is slow and resource intensive. Here, we present a high-throughput screening platform, coupled with a design-of-experiments (DoE) strategy, to identify optimal LC/HC expression balance at the transient stage. The system uses single-vector constructs encoding both LC and HC under promoters of defined low, medium, or high strength, enabling combinatorial testing of LC/HC promoter pairs. We applied this workflow to three different antibodies and quantified titer, viable cell density, and viability 72 h post-transfection. The optimal LC/HC promoter ratio was antibody specific. For two antibodies, high LC combined with medium HC expression yielded the highest titer while maintaining cell viability. High LC with high HC expression (LC100–HC100) also produced high titer but caused reduced viable cell density and viability. For the third antibody, the best-performing configuration was medium LC with medium HC, with medium LC and high HC as a close second. Across all three antibodies, low-strength promoters for either chain consistently resulted in poor titer. Overall, this platform offers a rapid and scalable approach to define antibody-specific LC/HC promoter strength combinations that maximize productivity without compromising cell health, enabling more informed construct selection before committing to stable clone generation.

Figures