Carbamazepine (CBZ) is indicated for the treatment of neurological disorders and can induce HLA-associated severe cutaneous adverse reactions (SCARs) in HLA-A*31:01+ patients. Positive predictive values for HLA-associated CBZ-induced SCARs indicate the involvement of further biomarkers despite the allele HLA-A*31:01. The aim was to further illuminate CBZ hypersensitivities with the objective to analyze differences in the TCR repertoire of HLA-A*31:01+ CBZ-responders vs. CBZ-non-responders.

To ensure absorption of CBZ in HLA-A*31:01+ PBMCs/DCs a mass spectrometric method using UPLC-MS/MS was performed. In an autologous setting a cytotoxicity assay was established using CBZ treated DCs as targets and CD8+ T cells from the same HLA-A-31:01+ donor as effectors. A Granzyme B ELISPOT assay has been utilized to detect reactive CD8+ T cells. Target cell viability has been assessed with annexin V and PI. This method enabled the characterization of HLA-A*31:01+ CBZ-responders and non-responders. To identify a potential public TCR phenotypical alterations of CD8+ T cells from classified donors were examined via RT-qPCR; a sophisticated method to sequence TCRs from HLA-A*31:01/CBZ reactive CD8+ T cells via NGS has been established.

An upregulation of TCR Vβ7 in CBZ responders, but not in CBZ non-responders could be observed. NGS analysis revealed significant alterations in the distribution of CDR3 length, clonotype abundances, HILL numbers, D50 diversity index and true diversity. TCRs from HLA-A*31:01 CBZ-responders vs. non-responders could be clearly distinguished.

We could significantly demonstrate that CBZ-reactive CD8+ T cells from HLA-A*31:01+ donors show a distinct TCR pattern and the existence of a public TCR could be identified. PCR analysis for the occurrence of CBZ-hypersensitivity additionally to HLA-typing should be implemented in patient management strategies prior to drug treatment. This would allow drug treatment of patients that are excluded from certain medications due to their genetic profile.

The authors declare no conflict of interest