CD44v6-targeted CAR T-cell therapy for head and neck squamous cell carcinoma

Introduction Immunotherapy with chimeric antigen receptors (CARs) is a future therapy option for head and neck squamous cell carcinoma (HNSCC). An artificial CAR construct consisting of a single chain fragment (scFv), linker, transmembrane and cytoplasmic domain combines tumor-associated antigen (Ag) recognition and T-cell activation. CD44v6, a splice form of CD44, is a suitable HNSCC-associated antigen. Therefore, we developed a CD44v6-specific CAR T-cell therapy against CD44v6⁺ HNSCC cells in vitro.

Method 33 primary HNSCC cell lines (CL) were screened for their CD44v6 and CD44 expression via flow cytometry. As a positive control CD44v6 cDNA was lentivirally introduced into the CL UT24A. The CD44v6 CAR construct was lentivirally expressed in primary human T-cells. CD44v6 CAR⁺ T-cells and HNSCC cell lines were co-cultured for 16h to assess the cytotoxic potential of the genetically modified T-cells. Lysis of HNSCC cells was determined by a CellTiter proliferation assay (Promega).

Results HNSCC CL show varying expressions of CD44v6 and CD44. Co-cultivation experiments of HNSCC cells with CD44v6 CAR⁺ T-cells revealed that the killing efficiency of the genetically modified T-cells is predominantly dependent on the Ag density on the target cells: CL with higher CD44v6 expression levels (UM14C, UM11B, UM10BPT) were efficiently lysed overnight already at low target-to-effector cell ratios, while UT24A cells with a low CD44v6 expression were less efficiently eradicated. Lentiviral overexpression of CD44v6 increased lysis of cell line UT24A by CD44v6-targeted CAR T-cells, indicating the pronounced specificity of the CD44v6 CAR.

Conclusion In this proof-of-concept study we established a specific CD44v6-targeted CAR T cell therapy for HNSCC.

Poster-PDF A-1296.PDF

Publication History

Article published online:
10 June 2020