With a market size of over $400B, biologics occupy the largest share of the total protein manufacturing market. However, the manufacturing effort is estimated to cost $80B annually, with production costs ultimately passed on to patients. Difficult-to-express proteins, expensive processes, low-yielding cell lines, challenges with maintaining glycosylation bio-fidelity, and inefficient scale-up pose significant hurdles to cost- effective and scalable protein production. Current solutions, including codon optimization and predictive AI-based technologies, have so far failed to universally facilitate high-yielding, low-cost, and scalable biomanufacturing. We have discovered a novel translational regulatory mechanism by which cells adapt to stressors by reprogramming the levels of tRNAs and over two-dozen chemical modifications of tRNA to enhance the translation of stress response proteins needed for survival of the stress at hand. Codomax has developed the Epi-MAX platform, an integrated omics and computational based platform that leverages this mechanism to enable enhanced translation of difficult-to-express proteins via bespoke tRNA-centric codon and cell line engineering in response to the stresses associated production of a target protein. The initial iteration of the platform increases recombinant protein expression by engineering codon usage in the target gene to match the stress-reprogrammed tRNAs, which to date has yielded up to 25-fold boosts in production of difficult-to-express proteins in Pichia, benchmarked against current codon optimization and strain engineering technologies. Despite the utility of Pichia as one of the major hosts for industrial protein manufacturing, protein production is plagued by intracellular aggregation and faulty transport through the secretory pathway. We have now extended the platform’s capabilities to increase stress translation of endogenous chaperones that prevent intracellular protein aggregation and facilitate transport of secreted recombinant proteins. Here, we will highlight application of the Epi-MAX platform’s tRNA-driven engineering approach to increase both translation and assembly of a panel of recombinant biologics in Pichia via translation engineering of chaperones, tRNAs, and tRNA modification proteins. These data highlight a novel epitranscriptomics-driven strain engineering approach to achieve the efficient, cost-effective, and scalable production of difficult-to-express biologics using Pichia.

Stephen Fried

Johns Hopkins University

Appearances