W171 - Gamma Delta T-cells Arming Bispecific Antibodies: Application to Neuroblastoma

Abstract Text: γδ T-cells are alternative cytotoxic effectors that recognize tumors through TCR and innate receptors, with limited lifespan that can be offset with multiple doses from clinical-grade expansion. In this context, bispecific antibodies (BsAb) arming T-cells are a viable and safer alternative to CAR transduction. Targeting neuroblastoma through O-acetyl-GD2 (OAcGD2), a GD2 variant with higher cancer-specificity, or B7-H3, a cancer-antigen with a broader expression and immunosuppressive roles, may further enhance efficacy.
We aim to propose alternatives to GD2 immunotherapy through production B7-H3 & OAcGD2/Vγ9Vδ2T-cells BsAbs and assessment of their functionality against neuroblastoma.
Using the Expi293 system, two BsAbs were produced containing an anti-Vδ2 nanobody linked to anti-OAcGD2 (B6D7) or B7-H3 (376D7) single-chain variable fragment. Purification involved 6xHis-tag binding on cobalt columns. Through flow cytometry, EC50 was assessed for BsAbs characteristics: binding to targets, induction of Vγ9Vδ2T-cells degranulation, and anti-neuroblastoma cytotoxicity at different effector: target (E:T) ratios.
Binding of both construct (B6D7 & 376D7) was observed on the OAcGD2+/B7-H3+ LAN-1 cells, but absent on OAcGD2-/B7-H3- Neuro2a cells. Similarly, equivalent binding on γδT-cells was detected for 376D7 and B6D7. Both constructs were able to induce degranulation of Vγ9Vδ2T-cells against antigen positive (OAcGD2 or B7-H3) cells. Furthermore, both constructs triggered cytotoxicity against antigen positive cells, with the percentage of apoptotic tumoral cells increasing with BsAb dose and E:T ratio
In conclusion, δT-cells BsAb can be produced and purified, and are functional. Ongoing work includes assays accounting for recently identified neuroblastoma cell identity diversity and constructs incorporating IL-15.