Metabolic regulation of T cell effector function and anti-tumor immunotherapy

Division Of Basic Sciences - Nci

Investigators

Linked publications & trials

Abstract

Despite significant advances in anti-tumor therapies, metastatic cancers remain challenging to treat. Adoptive immunotherapies-particularly those involving the infusion of ex vivo-manipulated, anti-tumor T cells-continue to show strong therapeutic potential. However, enhancing the persistence and functional responsiveness of these adoptively transferred cells remains a critical focus. The function of T cells engineered to express tumor-specific T cell receptors (TCRs) or chimeric antigen receptors (CARs) is often impaired by the hostile metabolic tumor microenvironment, which is shaped by nutrient availability, waste accumulation, oxygen concentration, pH, and physical forces. These distinct metabolic conditions may partly explain the success of CAR T cells in hematologic malignancies (e.g., CD19 CAR T cells for diffuse leukemias) and their limited efficacy in solid tumors. To overcome this barrier, we are focusing on the engineering of metabolically resilient CAR T cells via nutrient transporter modification and pharmacological targeting of key metabolic enzymes. Our goal is to improve T cell persistence and function in diverse tumor microenvironments. We hypothesize that CAR T cell therapeutic potential is influenced by differences in manufacturing platforms and can be predicted by measuring antigen-dependent functional activity. In collaboration with Dr. Steven Highfill (CPS, CCR), we have demonstrated that both T cell selection processes and the clinical manufacturing protocol significantly affect cytokine secretion profiles and in vitro tumor cell killing capacity-highlighting the importance of manufacturing parameters in the development of future clinical trials. To further characterize dynamic metabolic and functional properties of CAR T cells, we are collaborating with Dr. Grégoire Altan-Bonnet's laboratory (LICI, CCR) to leverage the high-throughput IMMUNOtron system. IMMUNOtron enables multiplexed, time-resolved analysis of CAR T cells by capturing over 10e7 data points-including 50 cytokines and surface markers across 12 time points. Using this computational platform, we have monitored the real-time behavior of CAR T cells from patients treated with CD22-CAR and CD19/CD22-CAR constructs at the NIH (in collaboration with Dr. Nirali Shah, POB) and identified significant differences compared to CD22-CAR T cells manufactured using a distinct platform at Stanford University (PIs: S. Ramakrishna and C. Mackall). Our data clearly demonstrate that manufacturing methodology profoundly impacts CAR T cell phenotype and function. This is among the first studies to assess how manufacturing platform differences influence patient outcomes resulting from treatment with T cells expressing an identical CAR-specifically, the CD22 CAR. Building on insights from Dr. Shah's Phase I trial of the bivalent CD19.22.BBz CAR, we integrated IMMUNOtron-based analyses to design a novel bicistronic CD19.28z/CD22.BBz construct with enhanced dual-targeting efficacy. This construct has progressed to evaluation in a Phase I/II clinical trial for pediatric B-ALL (NCT05442515; PI: N. Shah). Additionally, given the high frequency of CRLF2 rearrangements in high-risk pediatric B-ALL, we developed and submitted an Investigational New Drug (IND) application for a novel CAR T cell targeting the CRLF2 product, thymic stromal lymphopoietin receptor (TSLPR). The application was approved by the FDA in 2025. We are also developing a new generation of CAR constructs by engineering variations in the hinge domain, guided by molecular dynamics modeling in collaboration with Drs. Pratyush Tiwary (University of Maryland) and Grégoire Altan-Bonnet (NCI). In a separate collaboration with Dr. Paul François (University of Montreal) and Dr. Altan-Bonnet, we discovered that CAR T cell activation is enhanced by strong TCR antigen engagement, whereas weak self-antigen stimulation suppresses CAR activity. By leveraging this intrinsic property of TCR signaling, we designed a novel Antagonism-Enforced Braking System (AEBS) for CAR T cells. AEBS-CAR T cells exhibit robust tumor cytotoxicity while minimizing damage to healthy tissues, effectively reducing on-target/off-tumor toxicity. Harnessing this pre-existing inhibitory crosstalk between immune receptors opens new avenues for the development of more precise and safer cancer immunotherapies.