We began the interview by talking about how increasing monoclonal antibody yield has been a focus of bioprocessing scientists for decades, but more recently, product quality attributes, such as glycosylation, have increasingly become an area of focus. Dr. Elhofy explained that scientists realized that post-translational modifications to the protein were having an effect on the way the protein behaved. For instance immunogenicity, drug half-life, binding capacity, etc. is affected by post-translational modifications and impacts the effectiveness of the therapeutic. So to understand these post-translational modifications and to be able to impact them is a powerful tool in manufacturing therapeutics. Similarly, biosimilar products need to meet the pre-established parameters for post-translational modifications in order to have similar performance as the originator drug.
Next we discussed how cell culture optimization has also evolved from just looking to improve titer to now also optimizing for protein quality. I asked Adam if he could share some of the challenges associated with optimization efforts aimed at manipulating titer and quality. He stated that original stable clone screening was based on selecting for titer, then production, and finally post-translational modifications. These screening steps are now often happening simultaneously. Also, we now know that optimizing for higher titer can affect glycosylation. Adam gave the example of pH range, which impacts glycosylation patterns. Lower pH impacts more lower order glycoforms, and in some cases, the higher titer cultures are running at lower pH. Conversely, sugar nucleotides can be used to change glycosylation profiles, but can also cause reduced proliferation and titer. So often what improves titer is in direct conflict with tools for adjusting glycosylation patterns. Thus optimization schemes are becoming more complex.
I then asked if there was a way to optimize titer and glycosylation that had not been fully explored yet. To answer my question, Adam provided a terrific explanation of the cell biology related to glycosylation. He described how and where glycosylation occurs in the endoplasmic reticulum (ER) and Golgi and ways to improve function. These are both lipid membranes involved in the secretion and post-translational modification of proteins, however they are largely ignored in optimization strategies.
This was not always the case, there was research to support this approach. People tried adding lipids and cholesterol to boost the function of the ER and golgi, however these attempts weren’t successful. These failed attempts were primarily due to the lipids not being soluble in media. As a result, the addition of lipids caused problems in production including clogged filters, lipids that oxidized then became toxic and lipids adhering to vessel walls.
At Bio-Ess Laboratories, they looked for a way to solve this problem and created Cell-Ess. Cell-Ess is a novel technology that adds cholesterol and lipids to the media in a way where they stay soluble and are stable. The addition of Cell-Ess improves the energetics of the system and aids in the formation of the membranes of the ER and the Golgi. The ER is a low cholesterol organelle and the golgi is a high cholesterol organelle. If there is no cholesterol in the media, then understandably, there is going to be some stress to the system. The ER is also where ribosomes are located and they are responsible for the translation of MRNA into protein, so if the ER is under stress, there will also be a negative effect on titer. This is also where post-translational modification occurs and in the golgi is where you find the higher order glycoforms.
By adding Cell-Ess, you can change the dynamics of the glycosylation and improve the consistency of the glycolytic pattern. It has also been shown to aid in the addition of more higher order glycoforms. Adam describes Cell-Ess as another lever to improve titer and get the targeted post-translational modifications.
Next I asked about the challenge of adding cholesterol and lipids to media and whether Cell-Ess creates the same issues. Adam said when they created Cell-Ess they were aware of the issues with the current technology, and in response, they created an entirely different delivery mechanism for lipids and cholesterol. In this design, they ensured that Cell-Ess would be stable at 4 degrees and would not oxidize or create toxicity. Since it behaves similarly to cell membranes, it doesn’t adhere to vessel walls, just like a cell wouldn’t. Purification testing has been done and the cultures with Cell-Ess do not perform differently than control in terms of purification efficiency or filter clogging.
I then followed up by asking if there were any obstacles to adding Cell-Ess that needed to be addressed. He said that during product development they asked about the optimal way to bring Cell-Ess into applications. Scientists responded that they didn’t want to be required to change their media, feed structure, or timing. They wanted to be able to just add Cell-Ess to their current system, so that is how the product is designed. Cell-Ess is also manufactured under cGMP conditions, has a drug master file, is animal-component free and chemically defined. So Bio-Ess Laboratories has eliminated several of the implementation concerns up front.
To close the interview, I asked Adam if there was anything else he wanted to share with listeners. Adam wanted to highlight that with sequential optimization schemes, people often have to make a devil’s choice. If they want to fine tune their glycosylation and add higher order glycoforms, they often have to sacrifice titer and the opposite is true as well. With Cell-Ess they have worked to create a novel optimization tool to eliminate the devil’s choice problem and positively impact both titer and glycosylation pattern by boosting performance of the ER and Golgi.
To learn more about how Cell-Ess enables increased titer and improved quality, visit www.cell-ess.com.
Dr. Elhofy developed the core technology for the Ess line of products and aided in creating patents around novel uses of materials. Dr. Elhofy has over 14 years of scientific research experience in the areas of immunology, neuroscience, and oncology. He was funded by both the National Institutes of Health and the Multiple Sclerosis society as an investigator at Northwestern University Medical School. His doctoral research won him the award of the Top 5 trainee scientists by the American Association of Immunologists. Dr. Elhofy has 14 scientific publications in peer reviewed journals. He has played a variety of roles with start-up biotech companies ranging from Principal Investigator to Director of Corporate Development.