Genetic engineering of T-cells for the effective treatment of cancer

CD8+ T-cells can eliminate cancerous cells in vivo; and promising results have been documented on administration of in vitro expanded CD8+ T-cells. This has sparked great interest in the use of cellular therapy against cancer. However, "natural" anti-cancer T-cells are not very antigen sensitive and as such, not very effective at killing tumour cells. One of the most common reasons for these observations is that cancer-specific T-cell receptors (TCRs) are characterized by very weak affinities as high affinity TCRs are deleted during negative selection in the thymus. To overcome this challenge, a synthetic biology approach will be used to engineer novel T-cells that can efficiently kill cancer cells expressing a cancer antigen called '5T4'.
Aim 1: Identification of high affinity TCRs for 5T4, using an alloreactive approach TCR repertoires that have not been subjected to negative selection and are not HLA-class matched TCRs with high affinity for 5T4 will be sequenced and expressed in primary CD8+ T-cells.
Aim 2: Design CD8 variants that afford optimal recognition of 5T4: Computational modelling will be used to design novel CD8 variants which will be tested using biophysical analysis. CD8 variants will be expressed with a range of wild type and 5T4 specific TCRs.
Aim 3: Validation of genetically engineered T-cells: The ability of genetically engineered CD8+ T-cells to kill 5T4 positive tumour cells will be validated using 5T4+ mesothelioma cell lines and 3D mesothelioma spherocyte cultures which are representative of the tumour microenvironment.