Periodic Reporting for period 4 - VDJtargeting (Engineering T cells and B cells for Immunotherapy using V(D)J recombination)
Periodo di rendicontazione: 2022-04-01 al 2022-09-30
T cell engineering for immunotherapy has already saved many lives, but it cannot be applied to most patients in need, because it relies on cumbersome and expensive manipulations that can be performed in specialized centers only. In addition, B cell engineering has not shown therapeutic efficacy to date. Here, we develop a novel immunotherapy approach, specifically targeting immune genes into the genome of developing immune cells. Only such developing cells can incorporate the desired receptor or antibody gene. Therefore, only the most young and potent cells are engineered and later selectively expand when fighting their target, for example: the cancer cells. In addition, our engineered cells retain immune memory and thus reduce the risk of relapse. Our approach may revolutionize immunotherapy as it allows safe, efficient and scalable engineering of B cells and T cells, both outside and inside the body.
We made important proofs of concept in cell lines and in primary cells of human and mouse, inside and outside of the animal's body, in T cells and in B cells. Using injections into the thymus, we demonstrated gene integration into breaks occurring during T cell development. In addition, we demonstrated the expression of a desired T cell receptor following viral targeting of developing human T cells from cord blood. In B cells, we demonstrated antibody gene integration in a developing cell line. We further demonstrated antibody gene integration into breaks induced during B cell activation leading to secretion of large amounts of the desired antibody. When using CRISPR/Cas9 we further showed that upon immunization of mice, engineered B cells predominate over the natural response and differentiate into memory and antibody secreting cells while undergoing class switch recombination. A second immunization increases the activation of engineered B cells, indicating memory retention. Importantly, antibody sequences of engineered B cells in the spleen show evidence of evolution to counteract viral escape. Finally, we are the first to show genome editing of both T cells and B cells inside the animal's body. T cells and B cells may thus be engineered in a safe and potent manner as living and evolving drugs.
We have provided an extensive toolbox of novel and unconventional technologies for the editing of immune cells:
1. Injections into the thymus in order to engineer developing T cells
2. Engineering of developing T cells from the cord blood
3. T cell genome editing using breaks that occur naturally during cell development
4. B cell genome editing using breaks that occur naturally during cell development and during cell activation
5. B cell engineering using CRISPR to allow antigen induced activation, class switch recombination, and evolution to counteract viral escape
6. Genome editing of T cells and/or B cells using systemic injections
1. Injections into the thymus in order to engineer developing T cells
2. Engineering of developing T cells from the cord blood
3. T cell genome editing using breaks that occur naturally during cell development
4. B cell genome editing using breaks that occur naturally during cell development and during cell activation
5. B cell engineering using CRISPR to allow antigen induced activation, class switch recombination, and evolution to counteract viral escape
6. Genome editing of T cells and/or B cells using systemic injections