经 89Zirconium Oxine 放射性标记用于 PET 成像的表达 scFv-IL-13Rα2 嵌合抗原受体修饰 T 细胞的表征

Background Chimeric antigen receptor (CAR) T cell therapy is an exciting cell-based cancer immunotherapy. Unfortunately, CAR-T cell therapy is associated with serious toxicities such as cytokine release syndrome (CRS) and neurotoxicity. The mechanism of these serious adverse events (SAEs) and how homing, distribution and retention of CAR-T cells contribute to toxicities is not fully understood.

Methods To determine if radiolabelling of CAR-T cells could support positron emission tomography (PET)-based biodistribution studies, we labeled IL-13R 2 targeting scFv-IL-13R 2-CAR-T cells (CAR-T cells) with 89 Zirconium-oxine ( 89 Zr-oxine), and characterized and compared their product attributes with non-labeled CAR-T cells. The 89 Zr-oxine labeling conditions were optimized for incubation time, temperature, and use of serum for labeling.

In addition, product attributes of radiolabeled CAR-T cells were studied to assess their overall quality including cell viability, proliferation, phenotype markers of T-cell activation and exhaustion, cytolytic activity and release of interferon- upon co-culture with IL-13R 2 expressing glioma cells.

Results We observed that radiolabeling of CAR-T cells with 89 Zr-oxine is quick, efficient, and radioactivity is retained in the cells for at least 8 days with minimal loss. Also, viability of radiolabeled CAR-T cells was similar to that of unlabeled cells as determined by TUNEL assay and caspase 3/7 enzyme activity assay.

Moreover, there were no significant changes in T cell activation (CD24, CD44, CD69 and IFN- ) or T cell exhaustion(PD-1, LAG-3 and TIM3) markers expression between radiolabeled and unlabeled CAR-T cells. In chemotaxis assays, migratory capability of radiolabeled CAR-T cells to IL-13R 2Fc was similar to that of non-labeled cells.

Conclusions Importantly, radiolabeling has minimal impact on biological product attributes including potency of CAR-T cells towards IL-13R 2 positive tumor cells but not IL-13R 2 negative cells as measured by cytolytic activity and release of IFN- .

Thus, IL-13R 2 targeting CAR-T cells radiolabeled with 89 Zr-oxine retain critical product attributes and suggest 89 Zr-oxine radiolabeling of CAR-T cells may facilitate biodistribution and tissue trafficking studies in vivo using PET.