How to assess the potency and functionality of advanced therapies using cell-based bioassays
Gene and cell therapies are transforming the treatment of diseases for which therapeutic options were very limited until just a few years ago. These medicinal products offer new possibilities, but they also pose significant challenges throughout their development, manufacturing, and quality control.
One of the main challenges is demonstrating that a product not only contains the expected components but can also perform the biological function for which it was designed. Cell-based bioassays are essential tools for this purpose, as they enable the evaluation of biological activity and the determination of the potency of these medicinal products.
In this article, we explain what cell-based bioassays are, how they are applied to gene and cell therapies, and the main requirements for developing reliable and reproducible methods.
What are cell-based bioassays and why are they important?
Cell-based bioassays are experimental methods that use living cells to measure a biological response triggered by a product. Unlike conventional physicochemical analyses, they allow functional processes to be studied, such as receptor activation, protein expression, cell proliferation, or the destruction of target cells.
These assays are particularly relevant for gene and cell therapies because their mechanisms of action may involve several biological processes. Therefore, analyzing only the composition or quantity of an active substance does not always make it possible to determine whether the medicinal product retains its functionality.
For example, a gene therapy intended to correct an enzyme deficiency must demonstrate that the administered genetic material enables the production of a functional protein. In CAR-T cell therapy, which is designed to recognize specific tumor antigens, the ability of modified lymphocytes to eliminate cells expressing that antigen can be assessed.
These examples illustrate why cell-based bioassays are essential for characterizing the functionality of advanced therapy medicinal products.
Biological activity and potency in gene and cell therapies
Although closely related, the concepts of biological activity and potency are not equivalent.
Biological activity describes a product’s ability to trigger a specific response in an experimental system. Potency, on the other hand, is a quantitative measure of that activity, related to the biological attributes relevant to the medicinal product’s mechanism of action.
During the development of gene and cell therapies, potency testing provides information about product functionality and helps establish controls to ensure batch-to-batch consistency.
Regulatory recommendations from the European Medicines Agency (EMA) and the U.S. Food and Drug Administration (FDA) emphasize the importance of having appropriate potency methods in place to demonstrate relevant biological activity.
This requires selecting assays capable of detecting significant differences between batches and identifying changes that could affect the quality of the medicinal product.
It is important to remember that a positive bioassay result does not, on its own, demonstrate clinical efficacy. However, it provides essential information about product functionality and consistency throughout development and manufacturing.
Main bioassays used in gene therapies
In gene therapies, the methods employed depend on the vector used, the target tissue, and the intended mechanism of action.
Key analytical approaches include:
- Transduction assays: Evaluate the ability of a viral vector to deliver genetic material into cells.
- Gene expression assays: Quantify the production of RNA or proteins derived from the therapeutic gene.
- Enzymatic activity assays: Determine whether the expressed protein retains its expected biological function.
- Functional assays: Analyze whether the genetic modification produces the intended biological response.
For example, in gene therapies based on adeno-associated virus (AAV) vectors, measuring only the number of viral particles is not sufficient to fully establish functional potency. Methods that assess relevant aspects of product activity must also be incorporated.
Combining complementary assays can provide a more comprehensive characterization, provided that their selection is justified by the mechanism of action and the specific characteristics of the medicinal product.
How to assess potency in cell therapies
Cell therapies present an additional layer of complexity: the medicinal product itself consists of living cells whose characteristics and activity may vary.
In CAR-T treatments, cytotoxicity assays can measure the elimination of target tumor cells. Other parameters, such as cytokine secretion, cell proliferation, and cell activation, can also be assessed.
For products based on mesenchymal stromal cells, assays related to their immunomodulatory activity may be used, provided there is a scientific rationale linking these assays to the proposed mechanism of action.
Method selection must take into account cell identity, biological variability, experimental conditions, and the specific characteristics of the product.
Furthermore, some cell therapies require a combination of complementary assays to adequately characterize potency and cover different aspects of their biological activity.
Validation of cell-based bioassays and regulatory requirements
For a cell-based bioassay to be useful in quality control, it must demonstrate adequate analytical performance for its intended purpose.
Relevant parameters may include precision, accuracy, specificity, measurement range, and robustness, depending on the characteristics of the procedure and its intended use.
It is also essential to control potential sources of variability, such as the cells used, reagents, culture conditions, and reference materials.
The ICH Q2(R2) guideline on the validation of analytical procedures and ICH Q14 on analytical procedure development provide relevant principles for establishing suitable, scientifically justified methods.
For gene and cell therapies, the potency control strategy must be tailored to the product’s characteristics and the available knowledge of its mechanism of action. This strategy may evolve as new information becomes available during pharmaceutical development.
How to improve the reproducibility of bioassays
One of the main challenges in developing gene and cell therapies is turning complex experimental procedures into reproducible methods that can be used routinely.
Protocol standardization, qualification of cellular materials, and the establishment of acceptance criteria help reduce experimental variability.
In addition, automated technologies and multiparametric analysis systems offer new opportunities to characterize cellular responses and improve the consistency of results.
Selecting an appropriate method should begin with a fundamental question: What biological response provides meaningful evidence of the product’s functionality? From there, it is possible to design an analytical strategy that links the mechanism of action to a measurable response.
Cell-based bioassays are essential tools for evaluating the biological activity and potency of gene and cell therapies. Their appropriate selection and validation make it possible to characterize medicinal product functionality, support batch-to-batch consistency, and strengthen the quality control strategy.
As these treatments progress toward more complex stages of development and manufacturing, robust, reproducible, and scientifically justified methods are essential to demonstrate that the product retains its expected biological properties.
Frequently Asked Questions
- What are gene and cell therapies? Gene therapies use genetic material to modify, replace, or introduce biological functions into cells. Cell therapies use cells as part of a treatment; these cells may be administered without modification or following modification and expansion processes. The two categories may overlap in certain medicinal products.
- What are cell-based bioassays used for in advanced therapies? Cell-based bioassays measure the biological response triggered by a medicinal product using living cells. They are used to study product functionality and contribute to potency assessment and quality control.
- What is the difference between biological activity and potency? Biological activity is a product’s ability to produce a specific biological response. Potency quantifies that activity using an appropriate method and makes it possible to assess whether the product maintains the expected level of functionality.
- How is gene therapy potency assessed? Potency can be assessed using transduction assays, gene expression assays, enzymatic activity assays, or functional response assays. The selection depends on the vector, the mechanism of action, and the characteristics of the product.
- Which bioassays are used in CAR-T therapies? Common methods include cytotoxicity assays against target cells and analyses of cytokine secretion, cell activation, and cell proliferation. The combination of methods depends on the mechanism of action and the quality control strategy.
