Generation of novel chimeric antigen receptors to enhance the specificity and activity of T cells for the adoptive immunotherapy for cancer

© 2014 Dr. Connie DuongAdoptive immunotherapy is a promising treatment for cancer, with response rates of up to 70% in metastatic melanoma. To broaden this approach, T cells have been genetically modified to express chimeric antigen receptors (CARs) to endow T cells with anti-tumour activity capable of recognising a range of different cancer types. This approach has shown encouraging results in recent clinical trials for the treatment of haematological malignancies, however it has shown only moderate activity against solid cancers. To date, only a small number of molecules involved in T cell signaling have been incorporated into CARs, resulting in their suboptimal activity. Therefore improvements in CARs are needed in order to realise the full potential of adoptively transferred T cells. We proposed that using multiple or alternate signaling domains could enhance CAR-mediated T cell function. In this thesis, we describe the use of a DNA library of signaling molecules to investigate novel combinations of signaling molecules that could mediate enhanced CAR activity in the Jurkat T cell line and primary human T cells. A novel single-chain variable receptor was discovered comprising DAP10, CD3ζ and CD27 signaling domains that was able to trigger enhanced T cell activity in vitro and in an adoptive transfer mouse model. Clinical trials utilising CAR modified T cells have in some cases resulted in resulted in severe autoimmunity due to T cell recognition of tumour-associated antigens expressed on normal tissues. It is anticipated that as the application and efficacy of adoptive immunotherapy increases, toxicity against normal tissue will become increasingly common. To address this, we proposed that a T cell will respond less against normal tissue if endowed with a tumour-associated antigen-specific activating CAR co-localised with a chimeric inhibitory receptor (CIR) that is capable of turning off the T cell following engagement of antigen on normal tissue. We generated several novel chimeric inhibitory receptors and demonstrated expression of both CAR and CIR in T cells, which were then characterised for function against tumour-associated and normal tissue antigen expressing cell lines. In conclusion, the combination of these novel chimeric receptors may lead to a more efficacious but safer therapy for cancer