Collaborating Investigator: Dr. Michel Sadelain
Affiliation: Director, Center for Cell Engineering; Professor, Immunology Program, Memorial Sloan Kettering

Funding Source: Stand Up to Cancer
Grant Number: n/a
Project Period: 07/01/07-06/30/20
Project Status: New

Significance

This collaboration will enable us to translate our findings regarding immunometabolism to this novel treatment modality for cancer. Aim 1 seeks to exploit the robust molecular biology of CAR T cell generation and T cell genome editing to make these cells more potent effector cells. Aim 2 will define novel media to promote T cell persistence and memory. Aim 3 will harness our small molecule approach to regulating metabolism in order to enhance effector function and promote memory. By developing these three Aims for CAR T cells, we hope to not only further advance adoptive T cell therapy but also develop a tool box for metabolic reprogramming that can be exploited to enhance cancer immunotherapy as well as a vast array of immunologically engineered systems.

Approach

Metabolically reprogramming CAR T cells by genetic disruption of TSC2
The Sadelain Lab has recently developed an in vivo mouse model that enables the testing of the efficacy of various CAR T cell constructs. In brief, NSG mice (NOD.Cg-PrkdcscidII2rgtm1wji/SzJ) are inoculated with Pre-B ALL NALM6 cells that express FFLuc-GFP enabling the tracking of tumor growth by bioluminescence. By adoptively transferring sub-optimal doses of CAR T cells (where tumor growth begins to fail), they have successfully compared the potency and persistence of different CAR T cell constructs. In this aim, we will generate CAR T cells with a knock out of TSC2 using the CRISRP/Cas9 system. The Sadelain lab has already established this method for the targeted disruption of the TCR α chain in CAR T cells (1). Once gRNAs have been validated for TSC2 disruption, the Sadelain group will compare the ability of the TSC2KO CAR T cells to eradicate tumor when compared to the standard CAR T cell in the NALM6 “stress test” model. Concomitantly, our lab will interrogate the TSC2KO CAR T cells and the control CAR T cells in terms of their metabolic reprogramming using the parameters depicted in Aim 1. By knocking out TSC2 in CAR T cells, we seek to engineer more robust effector cells that are maximally metabolically reprogrammed for effector function and can subsequently pharmacologically converted into long-lived memory cells.

Generating metabolically fit, more persistent effector and memory T cells
Using the “stress test” model depicted by Zhao et al (2), we will determine the ability of metabolically defined media and SGK1 inhibition with JHU3201-01 to enhance persistence of CAR T cells. We predict that these strategies will markedly enhance the persistence and hence efficacy of CAR T cells in this model. Furthermore, we will test the ability of JHU3202-01 to enhance persistence using the TSC2KO T cells. Using the pharmaco-genetic approach we will initially reprogram the cells metabolically for maximal effector function then reprogram them for maximal persistence and memory generation.

Push-Pull:

This CP is relevant to all 3 Aims of TR&D 3. Aim 1 will initially employ TCR transgenic T cells but then using the molecular techniques and expertise of the Sadelain lab will be applied to CAR-T which will then be tested in their systems. Likewise, after initially optimizing the metabolically defined media, this will be tested with the CAR-T from the Sadelain lab and then then tested in their model systems (push). Similarly, the SGK1 small molecule inhibitors with be applied to CAR-T both ex vivo and in vivo to determine if they can enhance the efficacy of these cells in the Sadelain lab models (push). Interestingly, this collaboration will also promote the development of a genetic approach to deleting SGK1 in CAR-T using the Sadelain expertise and technology (pull).

Literature cited

(1) Eyquem J, Mansilla-Soto J, Odak A, Sadelain M. One-step generation of ‘Universal’ CAR T cells. Mol Ther, ASGCT Annual Meeting, 2016. (2) Zhao Z, Condomines M, van der Stegen SJ, Perna F, Kloss CC, Gunset G, Plotkin J, Sadelain M. Structural Design of Engineered Costimulation Determines Tumor Rejection Kinetics and Persistence of CAR T Cells. Cancer Cell, 2015.