Patel, Aysha; (2017) Development of T cell based therapeutic strategies for childhood cancer neuroblastoma. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

High risk neuroblastoma poses a significant clinical problem in paediatric oncology and new treatment strategies are needed. Antibody-derived bispecific T cell engagers (BiTEs) and chimeric antigen receptors (CARs) are novel treatment options that redirect a patient’s own T cells to recognise and eliminate tumour cells; both have demonstrated promise in clinical trials for haematological malignancies. In this study I explored an empirical approach to BiTE design in order to identify the optimal format for redirecting T cell cytotoxicity against neuroblastoma cells. GD2 was used as a target antigen, based on its high level of expression across neuroblastoma tumours and limited expression on healthy tissues. BiTEs were designed with different single chain variable fragments (scFv) to bind GD2 and CD3 (T cell) antigens. We demonstrated that a high affinity for GD2 was determinant of improved cytotoxicity of T cells against neuroblastoma cell lines and an optimal linker length between the two scFvs impacted tumour cell targeting. The secretion of interferon-γ and proliferation by activated T cells occurred in a CD3-specific and GD2-specific manner, confirming target specificity of the BiTEs. In a second strategy; as an attempt to reduce the on-target off-tumour toxicity of targeting GD2, novel antigen O-acetyl-GD2 was explored as an improved target antigen due to its restricted tumour expression pattern. The latter is a requirement when aiming to induce a persistent anti-tumour response with CAR T-cell therapy. An O-acetyl-GD2 specific CAR was generated which showed selective specificity to the O-acetylated form of GD2. Finally, as a pre-clinical approach to develop BiTE and CAR T cell therapy for neuroblastoma in vivo, pilot experiments were performed in a transgenic neuroblastoma mouse model which has co-expression of the ALKF1174L mutation and MYCN oncogene. This work indicated that this murine model appears suitable to develop T cell based immunotherapy into an effective therapeutic approach for neuroblastoma.

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