Researchers have developed a flow cytometry assay to detect anti-drug antibodies (ADAs) binding to CAR T cells in a dose-dependent manner. While CAR T Cell therapy has been highly effective as a cancer immunotherapy, it can lead to immune related adverse outcomes, including ADAs, which can impact efficacy.
Detecting Anti-Drug Antibodies
This method, published in ImmunoHorizons, demonstrates that flow cytometry can sensitively quantify ADA binding, assess functional impact, and support robust ADA assay development for CAR T therapies.
“The development of an assay able to detect and monitor the presence of ADAs against the CAR T cells can help to characterize the immune response as well as the CAR T product before administration to the patients, as well as during ongoing treatment, hopefully increasing the treatment’s success,” said Dr. Sara Morgado Garcia, a Senior Scientist at Labcorp, and senior author on the study.
Developing the Assay
To develop the assay, researchers used CAR T cells targeting receptor tyrosine kinase-like orphan receptor 1 (ROR1), which is of interest as a target for tumors that overexpress ROR1 such as leukemias, lymphomas, and solid tumors. ROR1 CAR T cells were incubated with increasing concentrations of ROR1-IgG, representing an ADA molecule in this research. The ability of ROR1-IgG to bind to both CD4 and CD8 ROR1 CAR T cells was assessed.
The flow cytometry assay developed could detect the ADA-like molecule binding to CAR T cells in a dose-dependent manner and showed that ADAs functionally interfered with CAR T cell activity. The researchers established a plate‑specific cut point using naïve donor serum to enable discrimination between background signal and true ADA reactivity.
Improving CAR T Cell Therapy
“We hope this work provides researchers and developers of CAR T cell therapies with a practical, sensitive, and reproducible flow cytometry–based approach to detect ADAs directly on intact CAR T cells, preserving the native conformation of the CAR and avoiding epitope masking, which is a main advantage,” shared Dr. Morgado Garcia
Long-term, the researchers hope this assay will help standardize ADA monitoring across CAR T platforms and accelerate the development of safe, more effective cell therapies. The research team plans to use this assay to support a wider range of CAR constructs and cell-based therapies.
Building on this foundation, Labcorp intends to offer this assay broadly to future clients developing CAR T and other cell‑based therapies and integrate this approach into their service portfolio to support the growing need for robust immunogenicity assessment.
