γδ T cell activation by bispecific antibodies

• Comparison of [(Her2)2 × Vγ9] tribody with other selective Vγ9 γδ T cell agonists.
• Superiority of [(Her2)2 × Vγ9] tribody in triggering γδ T cell-cytotoxicity.
• Negligible induction of γδ T cell death by tribody [(Her2)2 × Vγ9].
• [(Her2)2 × Vγ9] tribody is an attractive tool for γδ T cell-based immunotherapy.

Bispecific antibodies have been successfully introduced into clinical application. γδ T cells are of special interest for tumor immunotherapy, due to their recognition of pyrophosphates that are overproduced by many tumor cells resulting in HLA-nonrestricted tumor cell killing. Here we describe in detail a [(Her2)2 × Vγ9] tribody construct that targets human Vγ9 T cells to HER2-expressing tumor cells. The direct comparison with other selective Vγ9 T cell agonists including phosphoantigens and nitrogen-containing bisphosphonates revealed the superiority of the [(Her2)2 × Vγ9] tribody in triggering γδ T cell-mediated tumor cell killing with negligible induction of γδ T cell death. In contrast, phosphoantigens and bisphosphonates are potent inducers of γδ T cell proliferation but less efficient enhancers of γδ T cell-mediated tumor cell killing. Collectively, our data identify unique properties of a γδ T cell-targeting [(Her2)2 × Vγ9] tribody which make it an attractive candidate for clinical application in γδ T cell-based tumor immunotherapy.

A summary of the activation of Vγ9Vδ2 γδ T cells by bispecific antibodies in the defense against pancreatic ductal adenocarcinoma (PDAC) cells. (A) Cytotoxicity: stronger cytotoxic potential of initially activated γδ T cells (Vγ9Vδ2 γδ T cells within PBMC or freshly isolated γδ T cells, left side) against PDAC cells (tumor cells) is triggered by T-cell receptor (TCR)-dependent binding of bispecific antibody [(Her2)2 × Vγ9] as compared to [Her2 × CD3] or exogenous phosphoantigens (PAg, e.g., bromohydrin pyrophosphate, BrHPP) or nitrogen-containing bisphosphonate (n-BP). n-BPs inhibit Farnesyl Pyrophosphate Synthase, an enzyme of the eukaryotic isoprenoid pathway, which results in intracellular accumulation and release of the phosphoantigen isopentenyl pyrophosphate (IPP). Similar to cytotoxic activity of initially activated Vγ9Vδ2 γδ T cells, cytotoxicity of short-term cultured (Vγ9Vδ2) γδ T cell lines is also more efficiently enhanced by [(Her2)2 × Vγ9] than by PAg; in addition, however, the [Her2 × CD3] construct is also quite effective (right side). The differential effect of [Her2 × CD3] on initially activated versus short-term cultured γδ T cells is of importance when considering the potential application of the [Her2 × CD3] construct for in vivo activation versus adoptive transfer of γδ T cells. (B) Proliferation/cell death: the stimulation of γδ T cells within PBMC co-cultured with PDAC cells with PAg, n-BP or [(Her2)2 × Vγ9] results in an inhibition of γδ T cell proliferation. In short-term activated γδ T cells co-cultured with PDAC cells, PAg and n-BP trigger activation-induced cell death, in contrast to the [(Her2)2 × Vγ9] antibody construct.Figure optionsDownload as PowerPoint slide