A universally usable, CD34-derived, hinge domain for the treatment of head and neck cancer as well as solid and hematological cancers using CAR T-cell therapy

Background CAR T-cell therapy has revolutionized cancer treatment, especially for hematological diseases, and is also a focus of research for solid tumors. In CAR T-cell therapy, the extracellular variable single chain fragment of a monoclonal antibody is connected via a hinge domain to a transmembrane and intracellular T-cell activation motifs, thereby combining antigen recognition and T-cell activation in a single molecule. The aim of the work is to use the hinge to improve CAR T-cell stimulation or enable their targeted elimination or detection and enrichment.

Methods We established six hinge domains based on the truncated form of CD34, which contained the epitope for the CD34 antibody (Qbend-10). The fragments were cloned into lentiviral CAR constructs and the CAR-positive T-cells were enriched with CD34-specific microbeads via the MACS system (Miltenyi). After in vitro functionality measurements (FACS, cytotoxicity), the CARs with C6 hinge were tested in vivo in NOD-SCID-gamma mice.

Results A final construct of 99 amino acids (C6) was the best candidate for an efficient> 95% enrichment using the MACS system and additinally allows the direct detection of the CARs on the T-cells. The C6 hinge was functionally indistinguishable from the approved CD8α hinge both in vitro and in vivo.

Conclusion We have shown that our CD34 C6 hinge domain can be used for a wide variety of CARs and malignancies. It shows a high killing efficiency without mediating unspecific activation by target antigen-negative cells. This makes the new C6 hinge ideal for CARs in clinical applications, including head and neck cancer.

Düsseldorf School of Oncology

 

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Conflict of Interest

The author declares that there is no conflict of interest.

Publication History

Article published online:
24 May 2022