A key concern for those working in the demanding and dynamic field of drug development is the need for accessible, rapid, and accurate test formats for measurement and quantification of critical parameters – such as drug activity and immunogenicity – that still maintain a representative biological matrix and background.
In this interview, we speak with Dr. Michael Tovey, Chief Scientific Advisor of Svar Life Science, and inventor of the iLite® technology – a cleverly engineered cell-based assay system with a dual reporter gene readout that is designed so it can be developed for virtually any pharmaceutical target. He and the Svar Life Science team in France have been awarded over 20 international patents on the inventions underlying the iLite® technology. Here, Dr. Tovey describes how the team at Svar France is positioned to help researchers overcome the existing challenges with assay development; outlines the benefits of customized cell-based solutions that are adapted to specific research needs; and shares the exciting implications this versatile technology has for the future of drug development.
MT: Over the last 15 years – and particularly over the last 5 years or so – there has been an increased demand for cell-based assays due, at least in part, to the increasingly bespoke therapeutics that companies are developing to treat diseases with complex pathologies, such as cancer, autoimmune, and inflammatory diseases.
For example, viral vectors (such as adeno-associated viruses or AAVs) that encode a transgene can be designed either to replace a defective gene or to provide the protein encoded by the defective gene (such as FVIII & FIX) for the treatment of hemophilia. In this instance, cell-based assays are required to both quantify the activity of the protein encoded by the transgene and to quantify the transduction efficiency of the vector.
Although the development of such novel biotherapeutics has led to considerable progress in the treatment of disease, these therapies are often associated with the development of an immune response – both cellular and humeral – and notably the development of neutralizing antibodies. As the name suggests, these antibodies can neutralize the biological action of the drug. So, even fully human monoclonal antibodies can elicit an immune response due to the absence of tolerance to the protein. Therefore, cell-based assays are essential in order to quantify the effect of neutralizing antibodies on the activity of the drug or drug product.
In the past, there has been resistance to adopting cell-based assays due to the need to cultivate cells in the laboratory and the difficulty associated with the transfer of such assays from one site to another. These difficulties have since been resolved thanks to the development of assay-ready frozen cells that allow an analyst to take a vial of frozen cells from the freezer and run the assay immediately without any form of pretreatment. These cells also make it very easy to transfer vials to multiple sites during clinical trials, for example. In addition, the use of assay-ready frozen cells is, in fact, associated with a higher degree of reproducibility than when the same cell line is cultivated in vitro.
Assay-ready frozen cells [are] associated with a higher degree of reproducibility.
MT: When a client requests the development of an assay – usually for the quantification of the potency of a novel drug – their knowledge of how the chosen therapeutic agent works is often very limited. Therefore, we must establish, often for the first time, how the drug interacts with cellular pathways so that we can create an iLite® assay that reflects as closely as possible the mechanism of action (MoA) of the drug product. We are very fortunate to have an exceptionally gifted team of scientists led by Dr. Christophe Lallemand, many of whom have been with Svar Life Science since its inception, to carry out this work in our labs.
Once the optimal pathway to target has been decided, we prepare a range of molecular constructs required for the assay and test these constructs in a panel of different human cells. Then, once the optimal construct-cell line combination has been established, the cloning process can begin. Typically, several hundred clones are screened and ten or so of the most promising clones are characterized further for performance and stability.
When we finally select a single clone, a master cell bank (MCB) and a working cell bank (WCB) are created and, again, characterized for performance and stability. This means, if required, we can then take a single vial of the WCB and prepare numerous assay-ready frozen cells. After developing the cell line and selecting the final clone, we then establish the optimal assay conditions for the drug product prior and develop the potency assay, determining its analytical range and assay robustness.
MT: First and foremost, the considerable knowledge of our experts at Svar Life Science allows us to choose the optimal strategy for establishing fit-for-purpose cell lines that reflect the MoA of the chosen drug product as closely as possible. These customized cell lines are designed to provide maximum sensitivity, specificity, and dynamic range, which are essential for the development of a potency assay that fulfills the requirements of regulatory authorities. These cell lines are also fully characterized for stability and performance over a long period of time. In addition, our MCBs, WCBs, and assay-ready frozen cells are developed and subject to rigorous quality control to ensure consistent results over the lifespan of the drug product.
Most of the reporter-gene cell lines that we develop also contain a second different luciferase reporter-gene under the control of a constitutive promoter, in addition to the drug-responsive luciferase reporter-gene. This allows drug-induced activity to be normalized with respect to the constitutive expression of the second reporter-gene rendering the assay independent of cell number and allowing for neutralizing antibodies to be distinguished from non-specific serum matrix effects.
Both conventional and complex novel drug products can benefit from this technology, and the development of precision assays is fundamental for this.
Customized cell lines are designed to provide maximum sensitivity, specificity, and dynamic range.
MT: We are often dealing with highly innovative products, which have never been produced before. So, it really comes down to the limited knowledge of the – likely complex – MoA of the drug product. The challenge is in choosing the pathway that reflects the MoA as closely as possible and can provide the basis for a specific and robust potency assay.
As mentioned earlier, our considerable knowledge in the development of custom iLite® reporter-gene assays has helped considerably. We have made cell lines for numerous big pharma and biotech companies and has always been successful in developing a custom cell line with the desired characteristics. To achieve this, we often develop parallel approaches and then innovate. Such innovations include the development of novel chimeric transcription factors and/or novel chimeric receptors, the latter of which is currently in progress for a large pharma client of ours.
Advice for researchers looking to incorporate customized cell lines into their work
Start the process early on. Dr. Michael Tovey advises providing your industry partner with the maximum amount of information concerning the MoA of the drug, and initiate discussions concerning the development process as soon as possible.
MT: Complex therapeutics and therapeutic drug monitoring are two key areas where iLite® technology is currently showing value.
Given the development of increasingly complex biotherapeutics for the treatment of diseases with complex pathologies, iLite® reporter-gene assays are uniquely positioned to meet these challenges. This technology has already been used to develop potency assays for numerous complex antibody therapeutics, including tri-specific monoclonal antibodies, AAV-encoded transgenes, and neutralizing antibody assays for both wild-type AAV serotypes and recombinant viral vectors.
The iLite® reporter-gene assays are also playing an increasing role in therapeutic drug monitoring, for example, in gastroenterology, where patients have been treated with anti-TNF-alpha monoclonal antibodies and monitored for drug activity and drug-neutralizing antibodies at regular intervals during therapy in conjunction with clinical evaluation.
Find out more about how Svar Life Science is assisting researchers in their work: