Degradation Products of Tryptophan in Cell Culture Media: Contribution to Color and Toxicity.

Biomanufacturing processes may be optimized by storing cell culture media at room temperature, but this is currently limited by their instability and change in color upon long-term storage. This study demonstrates that one of the critical contributing factors toward media browning is tryptophan. LC-MS technology was utilized to identify tryptophan degradation products, which are likely formed primarily from oxidation reactions.

Lactoyl leucine and isoleucine are bioavailable alternatives for canonical amino acids in cell culture media.

Increasing demands for protein-based therapeutics such as monoclonal antibodies, fusion proteins, bispecific molecules, and antibody fragments require researchers to constantly find innovative solutions. To increase yields and decrease costs of next generation bioprocesses, highly concentrated cell culture media formulations are developed but often limited by the low solubility of amino acids such as tyrosine, cystine, leucine, and isoleucine, in particular at physiological pH. This study sought to investigate highly soluble and bioavailable derivatives of leucine and isoleucine that are applicable for fed-batch processes.

Reactivity and degradation products of tryptophan in solution and proteins.

Tryptophan is one of the essential mammalian amino acids and is thus a required component in human nutrition, animal feeds, and cell culture media. However, this aromatic amino acid is highly susceptible to oxidation and is known to degrade into multiple products during manufacturing, storage, and processing. Many physical and chemical processes contribute to the degradation of this compound, primarily via oxidation or cleavage of the highly reactive indole ring.

Vitamins in cell culture media: Stability and stabilization strategies.

Nowadays, chemically defined cell culture media (CCM) have replaced serum- and hydrolysate-based media that rely on complex ingredients, such as yeast extracts or peptones. Benefits include a significantly lower lot-to-lot variability, more efficient manufacturing by reduction to essential components, and the ability to exclude components that may negatively influence growth, viability, or productivity. Even though current chemically defined CCMs provide an excellent basis for various mammalian biotechnological processes, vitamin instabilities are known to be a key factor contributing to the variabilities still present in liquid CCM as well as to short storage times.

Impact of S-sulfocysteine on fragments and trisulfide bond linkages in monoclonal antibodies.

The quality of recombinant proteins such as monoclonal antibodies produced using Chinese hamster ovary cell-based mammalian systems is dependent on many factors, including cell line, process and cell culture media. Due to these factors, the generated product is heterogeneous and may have chemically-induced modifications or post-translational modifications that affect antibody stability, functionality and, in some cases, patient safety. This study demonstrates that S-sulfocysteine, a cysteine derivative, can increase the antibody specific productivity in different cell lines cultivated with different processes while minimizing trisulfide linkages in generated mAbs, mainly between heavy and light chain.

Antioxidant effect of thiazolidine molecules in cell culture media improves stability and performance.

The ability of cell culture media components to generate reactive species as well as their sensitivity to oxidative degradation, affects the overall stability of media and the behavior of cells cultured in vitro. This study investigates the influence of thiazolidine molecules, formed from the condensation between cysteine and alpha-ketoacids, on the stability of these complex mixtures and on the performance of cell culture processes aiming to produce therapeutically relevant monoclonal antibodies. Results presented in this study indicate that 2-methyl-1,3-thiazolidine-2,4-dicarboxylic acid and 2-(2-carboxyethyl)-1,3-thiazolidine-2,4-dicarboxylic acid, obtained by condensation of cysteine with pyruvate or alpha-ketoglutarate, respectively, are able to stabilize cell culture media formulations, in particular redox sensitive molecules like folic acid, thiamine, l-methionine (met) and l-tryptophan (trp).

S-Sulfocysteine simplifies fed-batch processes and increases the CHO specific productivity via anti-oxidant activity.

Industrial fed-batch cultivation of mammalian cells is used for the production of therapeutic proteins such as monoclonal antibodies. Besides medium ensuring initial growth, feeding is necessary to improve growth, viability and antibody production. Established processes include a slight acidic main feed and a separate alkaline feed containing l-tyrosine and l-cysteine.

Improvement and simplification of fed-batch bioprocesses with a highly soluble phosphotyrosine sodium salt.

Fed-batch culture bioprocesses are currently used predominantly for the production of recombinant proteins, especially monoclonal antibodies. In these cultures, concentrated feeds are added during cultivation to prevent nutrient depletion, thus extending the cellular growth phase and increasing product concentrations. One limitation in these bioprocesses arises from the low solubility or stability of some compounds at high concentrations, in particular amino acids.