Advanced development of composite gene delivery and CAR engineering systems

SUMMARY | Cellular immunotherapy such as chimeric antigen receptor (CAR) T cells involves engineering and
adoptive transfer of cells to directly target tumor cells in patient, and demonstrated clinical success. However,
current cell forms of cell therapies face multiple major hurdles, centering around efficient generation of potent,
specific and safe therapeutic immune cells. This R33 will address this significant problem, by developing
and rigorously validating two sets of versatile gene delivery and cell engineering toolkits, to ease the
therapeutic cell generation issue, and to offer a simple yet distinct approach to achieve high potency. Aim
1. Advanced development of MAJESTIC, a highly efficient composite gene delivery system. A vital part of
cellular immunotherapies is therapeutic cell generation. Current approaches, including lentiviral or g-retroviral
vectors, AAV, mRNA, DNA transposons, and genome editing such as CRISPR/Cas, all have their own
limitations. In the first part of this R33, we will perform advanced development and validation of MAJESTIC
technology (mRNA AAV-Sleeping-Beauty Joint Engineering of Stable Therapeutic Immune Cells). This system
can transduce diverse immune cell types with minimal cellular toxicity, leading to highly efficient and stable
therapeutic cargo delivery. Aim 2. Advanced development of synthetic fusion tails to enhance therapeutic
cell function. Despite success in CAR-T therapy in hematopoietic malignancies, major challenges still exist
such as tumor antigen loss, T cell exhaustion, T cell dysfunction, and poor in vivo persistence that hampered its
widespread clinical potential. We seek to develop and validate a distinct approach for cellular engineering to
add to the armamentarium of tools to enhance CAR immune cells against cancer. In the second part of this R33,
we will advance the development of TAILFUSE technology, a novel and unique CAR engineering approach by
cytoplasmic tail (CT) fusions, which reprograms CAR-T function and substantially enhanced in vivo anti-tumor
efficacy. We achieved proof-of-concept development of the technologies (R21/R61 equivalent). In this project
we will perform robust validation, optimization, extension and advanced development (R33). This R33 will mature
these versatile tools for gene delivery and synthetic cell engineering, including quantitative performance
measures, benchmarking, new capability extension, and validation of broader applications to cancer-related cell
types. Success of this R33 will lead to novel technologies that will bring new capabilities, with substantial
improvements over existing technologies. We anticipate wide-spread use of such technologies in laboratory
research and therapeutic development settings by the field to reach transformative impact.

Grant Number: 5R33CA281702-03
NIH Institute/Center: NIH
Principal Investigator: Sidi Chen