Dipeptides to enhance yield and viability from cell cultures

What is this patent about?

’178 is related to the field of recombinant protein production in animal cell cultures, specifically focusing on optimizing serum-free and peptone-free media. In commercial bioprocessing, the transition away from animal-derived components like serum and plant-based protein hydrolysates is desirable for safety and consistency, but often results in nutritional deficiencies that trigger apoptosis and reduce protein titers. The invention addresses the need for chemically defined media that can sustain high cell viability and productivity without the heterogeneity of traditional additives.

The underlying idea behind ’178 is that specific synthetic dipeptides can serve as highly soluble, stable nutrient sources that trigger a metabolic shift from cell proliferation to increased protein productivity. By utilizing dipeptides containing tyrosine and histidine, the invention overcomes the inherent solubility limits of individual amino acids like tyrosine, which often become depleted in high-density cultures. This targeted supplementation effectively mimics the beneficial effects of complex peptones while maintaining a chemically defined environment, allowing for extended culture durations and improved metabolic profiles.

The claims of ’178 focus on a method and a resulting cell culture for producing proteins in recombinantly engineered Chinese hamster ovary (CHO) cells using a two-phase process. The independent claims specifically protect the use of a serum-free defined production medium supplemented with at least one dipeptide selected from a specific group, including Tyr-His, Tyr-Lys, His-Gly, and Ala-His. These claims cover the application of these dipeptides to improve either the final protein titer or the overall viability of the cell culture compared to standard defined media.

In practice, the invention works by introducing these dipeptides during the production phase, either in the initial batch medium or as part of a concentrated feed. This supplementation prevents the depletion of critical amino acids and helps manage the culture's pH and lactate levels. By maintaining these parameters, the dipeptides delay the onset of programmed cell death, enabling the production phase to be extended from the industry-standard 11 days to as long as 15 days, which significantly increases the total amount of protein harvested.

This approach differs from prior methods that relied on complex, undefined plant hydrolysates or larger tri- and tetra-peptides which are more difficult to characterize. Unlike simple amino acid supplementation, which is limited by the low solubility of tyrosine, the use of tyrosine-containing dipeptides ensures a steady supply of nutrients without precipitation. Furthermore, the invention identifies that specific structural orientations are critical, as evidenced by the finding that while His-Gly improves performance, its sequence reversal, Gly-His, can actually have a negative impact on the culture.