Your source for answers to the complex challenges of cell and gene therapy development.
We recently spoke with Labcorp Drug Development team member Paul Byrne, Principal Scientist, Cell & Gene Therapy, Biopharmaceutical CMC Solutions. He shared an introduction to CMC, key analytical methods used in assessing the Critical Quality Attributes, challenges in developing these assays and his thoughts on the road ahead over the next 5-10 years.
Can you provide a quick introduction to CMC and why it’s important in the development of cell and gene therapies?
Chemistry Manufacturing and Control (CMC) addresses all aspects of the manufacture of cell and gene therapy molecules, facilities and equipment. More importantly, it also entails characterization, quality control, batch release and stability. It encompasses a broad range of analytical tests to ensure, among other things, that the process is safe, effective and that there’s consistency between batches.
Since these molecules will be administered to preclinical species and patients, it’s critical that the material is safe and effective. To do so, we must factor in what exactly we’re working with. Currently, we utilize a variety of complex molecules, such as viral vaccines, gene therapy vectors, plasmid, oligo, siRNA, CAR-T, and gene editing.
CMC is approached on a case by case basis, so the analytical strategies can be quite different
To approach the development of the CMC analytical testing strategy, we focus on assessing the Critical Quality Attributes (CQAs) for that particular molecule. We ask the following questions: 1) How much do we have, 2) Is it safe, 3) How potent is it (is it biologically active?) 4) Is it pure, 5) Can we confirm identity, and 6) Is it stable?
We take what we know about the molecule and use the answers to these CQA questions to develop an analytical testing strategy. These CMC packages can be quite challenging, depending on the
molecule, and generally include quite complex analytics from a broad range of techniques.
What are the key analytical methods that are involved in assessing the Critical Quality Attributes for cell and gene therapy molecules?
The typical Critical Quality Attributes of safety, purity, identity, quantity, potency and stability, which are generally applied to biological products also apply to cell and gene therapies. We’ve introduced the principles of CMC for developing the analytical strategies around these CQAs.
As far as safety is concerned, CQAs are fairly standard across most current cell and gene therapy products. For starting materials like the cell lines, which are used to manufacture gene therapy products and retroviral/lentiviral vectors, safety testing focuses on potential viral contamination.
For final products, viral contamination testing becomes more specific. For example, testing for replication competent rcAAV is required for AAV products.
In contrast, for cell therapy, if the retroviral/lentiviral vector was used for transduction, testing for replication competent retrovirus is required. Viral testing methods can be qPCR (direct) and/or cell-based assays.
In addition to viral safety evaluation, microbiological safety testing (e.g., sterility, endotoxin and
mycoplasma) is required. This testing often relies on compendial methods. For CAR T-cells, vector copy number (or the number of integration events) is also considered a safety evaluation.
Compared to the standard safety assessments, analytical strategies for purity can be quite different, depending on the product and process.
The most important CQAs relate to the quantity and potency of these CGT products. AAV-based gene therapies rely on digital droplet PCR and ELISA methods to quantify the genomic and capsid titers. However, cell-based therapies like CAR T-cells rely on flow cytometry to quantify the desired cells and determine the viability of the cells.
Although cell and gene therapy products have complex biological activities and different mechanisms of action, we have seen similar approaches to potency assessment for a variety of products. When working with more complex cell therapy products, we see key differences. Sometimes one assay is not sufficient to measure the product attributes for potency. Therefore, one alternative approach is to develop multiple complementary assays which measure different product attributes associated with potency.
What are the challenges and considerations for developing and validating these assays?
There’s quite a few challenges and considerations when developing CMC assays. The first main challenge is how much validation to conduct at the different stages of product development. The current regulatory guidance recommends that assays do not need to be fully validated until the end of Phase III clinical or market approval as part of BLA/NDA submission. We take steps before that time to that to show that assays are suitable. There are three main phases in this regard: fit for purpose, partial validation and full validation.
According to regulators, prior to the first-in-man (FIM) clinical studies, you should at least have assays that are suitable for use. However, there is a risk associated with this. If you wait until your product is nearing approval to find out that some of the parameters have failed, there will be delays in obtaining approval.
To help combat these issues in more critical and complex assays, some companies perform partial validation prior to FIM studies, then quickly continue on with robustness and full validation. There is a risk here in terms of time and cost. But as more and more products come to market, clients are more confident of their molecule’s success, and are happy to invest time and money to perform these full validations early in the product life cycle.
The other main challenge lies in how we actually develop and validate these assays. For the genomic titer, a lot of work needs to go into design and development for it to be compliant with the very strict precision criteria currently being recommended by regulators.
For potency assays, there are significant challenges for what is already a complex assay, and a significant amount of work is required to develop and optimize prior to any validation activities. It isn’t unusual to have a development and optimization period of more than 12 months, so this is the assay to work on before anything else.
The FDA has recently forced several companies to delay development plans due to concern over manufacturing and the tests being used to assess the strength for of their treatments. Please share your thoughts on this.
I believe these recent issues highlight the importance of both the CMC assays and a suitable analytical testing strategy. It’s important to ensure that assays are at an appropriate level of validation. To solve this problem and make sure that regulators are happy with the CMC data, there’s a few key things to focus on. First, we should do as much as we can, as soon as we can, to understand the molecule that we are working with. This will help ensure that our analytical testing strategy uses the most appropriate tools to address CQA questions.
It’s critical to know how the assay is validated. For more complex molecules, where our understanding may be limited, assays should be well developed and optimized. They should also be partially validated as early as possible in the product life cycle. In addition, it would be helpful to have more guidance in terms of how these critical assays should be designed and validated. It’s clear that as much care as possible should be taken when developing these analytical testing strategies.
What kind of developments do you expect to see over the next five to ten years?
Future developments are hard to predict given that this area is changing so rapidly. However, we do have some indications for the future. Five CAR-T cell products have currently been approved by the FDA. All of them are autologous cells, and all are for blood cancer treatment. I think we will see expansion to solid tumors and perhaps other diseases in the next few years.
We may also see the allogeneic CAR T-cell products go to market. In fact, multiple allogeneic CAR T-cell products have already moved to the clinical phase. For example, Allogene AlloCAR T™ PIPELINE and Cellectis TALEN® gene-edited CAR T-cells. The preliminary data from Cellectis Phase I study of U-CAR-T-22 demonstrates “the promise of the off-the-shelf allogeneic cell therapy to leapfrog the autologous CAR-T products.”
In addition to more CAR T-cells, other cell therapy products, such as TRC T-cells and NK/NKT cells, will also move to later phases of development in the next few years. New gene editing technologies will be used for the newer generations of cell therapy to archive better safety and efficacy.
At the same time, scientists are trying to couple viral vectors with gene editing technique to develop novel gene therapy, using viral vectors to package, deliver, and express CRISPR or transposon components for targeted in-vivo gene editing.
These developments will result in new and probably more complex cell and gene therapy products, which will certainly impact the CMC lot release testing for autologous cell therapy products, which is currently very challenging due to the high lot number, the limited testing material, and the quick turnaround time.
With the allogeneic cells replacing autologous cells, the product lot number for the same number of patients will significantly decrease. Sample size and stability will also increase.
Another helpful development concerns non-viral vectors used for cell and gene therapy manufacturing. The current replication competent retrovirus (RCR/RCL) testing will no longer be needed. This is excellent news since it was labor intensive, time consuming and costly.
Overall, considering all the novel cell and gene therapy products that have developed over the last decade, I believe the future will be just as promising in our efforts to deliver safe and effective products that address patients’ unmet needs.
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