Improved Lentiviral Vectors for Immunotherapy

Abstract

A safe and efficient gene transfer for immunotherapy can be achieved through cell type-specific gene delivery using targeted lentiviral vectors (LVs). A promising technology for these LVs is based on the concept of the Adapter-LV, where measles virus (MV) envelope proteins are used for pseudotyping. This type of MV-LV uses a tag-specific single-chain variable fragment (scFV) fused to the viral attachment protein of MV, allowing flexible and selective transduction through tagged adapter molecules. However, MV-LVs have a low viral titer limiting their clinical applications. Additionally, MV-LVs require T cell activation for efficient transduction, but prolonged activation may also reduce the anti-leukemic potency of adoptively transferred T cells. This aspect emphasizes the importance of quiescent T cells as target population. This dissertation aimed to overcome these limitations by optimizing the Adapter-LV technology. To this end, an alternative pseudotype was assessed using the envelope proteins of the canine distemper virus (CDV). These CDV-pseudotyped LVs (CDV-LVs) were designed to achieve higher viral vector titer yields. Additionally, strategies to improve transduction efficiency of quiescent cells were explored and agonistic surface molecules were also employed on the CDV-LV to induce T cell activation. The results show that CDV-LVs exhibit significantly higher viral titer yields while maintaining the unique selectivity and flexibility of the foundational MV-Adapter-LV platform. Furthermore, these CDV-LVs successfully generated functionally active chimeric antigen receptor (CAR) T cells, as confirmed by various in vitro and in vivo tests. It was demonstrated that CDV-LVs enabled efficient transduction and activation of primary, non-activated T cells. In summary, using an alternative pseudotype improves gene transfer and can sustainably expand the application e.g. by incorporating additional molecules. Future work should assess the T cell activation required for CAR T cell generation by CDV-LVs.

Die Dissertation ist gesperrt bis zum 06. Februar 2028 !