Abstract
Process intensification and simplification in biopharmaceutical manufacturing have driven the exploration of advanced feeding strategies to improve culture performance and process consistency. Conventional media design strategies, however, are often constrained by the stability and solubility challenges of amino acids, particularly in large-scale applications. As a result, dipeptides have emerged as promising alternatives. Despite extensive research on amino acids, dipeptide supplementation in Chinese hamster ovary (CHO) cell-based manufacturing has received comparatively less attention. In this review, we critically analyze challenges associated with amino acids prone to instability and poor solubility (e.g., glutamine, cysteine, and tyrosine), and explore the potential of dipeptides to address these limitations. We explore the intricate mechanisms of dipeptide transport and enzymatic cleavage, highlighting how chemical properties, stereoisomerism, and competitive metabolites influence their utilization. Notably, while most dipeptides exhibit enhanced solubility, their stabilization effects and culture performance remain variable, underlining the need for rational design. To guide future innovations, we propose tailored dipeptide strategies derived for specific biomanufacturing needs by integrating multi-omics analysis, metabolic flux modeling, and artificial intelligence (AI) modeling. KEY POINTS : •Explored dipeptides as a solution to amino acid instability and poor solubility, enhancing cell culture performance. •Discussed transporter kinetics and cleavage enzymes influencing dipeptide utilization in biomanufacturing. •Suggested various design strategies for identifying appropriate dipeptide pairs to improve bioprocess efficiency.