Protransduzin™ - A novel transduction enhancer to accelerate en gros CAR-T production

Abstract

Background & Aim The advent of cellular immunotherapies using genetically reprogrammed immune cells such as T-lymphocytes with chimeric antigen receptor (CAR-Ts) or engineered T-cell receptors (TCR-Ts) has led to promising new treatments for hematological cancers. However, due to the complex multi-step manufacturing process, these therapies are still very costly and currently approved therapies exceed 400k US$ per patient. In order to make CAR-T immunotherapy more affordable, a reduction of the production-related cost of goods and process optimization is highly desirable. Methods, Results & Conclusion A crucial manufacturing step is the actual gene transfer of CARs and TCRs which to date mostly relies on lenti- or retroviral vectors and often results in low transduction rates. To date a range of genetic transfer enhancers is being used to improve transduction with variable success. These include amphiphilic macromolecules, cationic polymers, and adhesion molecules. Until now the coating of cultureware with recombinant fibronectin (or a fragment thereof) has been shown to provide fair enhancement and is widely accepted. However, coating procedure is cumbersome with time consuming liquid handling steps in a cleanroom environment. Here we describe Protransduzin™, a novel peptide that can enhance transduction of CARs in human leukapheresis-derived CD3 lymphocytes by up to 24% over control dependent on culture system and multiplicities of infection. A side-by-side comparison of Protransduzin™ with recombinant fibronectin and an additional, commercially available histidine-rich amphipathic peptide showed that Protransduzin™ is at least as potent as the adhesion molecule but more potent than the other peptide. Since Protransduzin™ can be added directly with the viral particles it can be used to streamline CAR-T production. Especially in connection with mini bio-reactors it is possible to do an en gros production by allowing transduction and expansion in one vessel. Thanks to our optimized protocol shown here hands-on time can be reduced along with the contamination risk arising from repeated liquid handling steps.