Abstract 3603: Reverse fate mapping of CD19-targeted CAR T cells in patients with large B-cell lymphoma
Authors: Good Zinaida , Hamilton Mark P. , Spiegel Jay Y. , Kurra Sreevidya , Desai Moksha , Prabhu Snehit , Yang Eric , Ozawa Michael G. , Hanson Paul J. , Wu Fang , Frank Matthew J. , Baird John H. , Muffly Lori , Claire Gursharan K. , Craig Juliana , Kong Katherine A. , Wagh Dhananjay , Coller John , Plevritis Sylvia K. , Sahaf Bita , Miklos David B. , Mackall Crystal L. 
Published: June 2022

Autologous T cells genetically engineered to express a chimeric antigen receptor targeting CD19 (CD19-CAR) have achieved high complete response rates in patients with hematologic malignancies, but &gt;50% of patients progress following therapy. Here, we sought to understand key T-cell intrinsic factors impacting efficacy, namely CAR T-cell expansion, persistence, and homing to the tumor. Using an approach called reverse fate mapping, we followed individual T-cell clones at the single-cell level from pre-manufacture apheresis to the infusion product, tumor-involved lymph node, and blood at peak and late expansion in 12 adult patients with relapsed or refractory large B-cell lymphoma treated with axicabtagene ciloleucel, an FDA-approved CD19-CAR T-cell immunotherapy. The resulting CAR T-cell atlas comprises matched transcriptome (scRNA-seq) and surface protein expression (CITE-seq) for 322,028 cells from 44 samples, with 119,397 unique T-cell receptor (TCR) clonotypes identified. This atlas enabled us to ask questions like: “What were the phenotypes of the most successful CAR T-cell clones at the time of infusion or pre-manufacture apheresis?” We found that T-cell clonotypes with juvenile features at apheresis, including IL7R expression, were the most successful at expansion to higher frequencies in the infusion product, while clones with effector gene expression programs, such as those encoding perforin and granzymes, contracted between apheresis and product. Conversely, it was GZMK-expressing T cells in pre-manufacture apheresis that were dominant in the tumor early following CAR T-cell infusion. Further, T-cell clonotypes with active effector programs at infusion dominated at peak expansion. Finally, we defined active expression modules and pathways in the infusion product for CAR T-cell clones that homed to the tumor or became dominant at late expansion. These analyses pinpoint the molecular mechanisms that could be modulated to rationally steer CAR T-cell differentiation trajectories at the genetic or pharmacological level. This work was supported in part by the Parker Institute for Cancer Immunotherapy, California Institute for Regenerative Medicine, Kite Pharma, and Stanford Cancer Institute.

Citation Format: Zinaida Good, Mark P. Hamilton, Jay Y. Spiegel, Sreevidya Kurra, Moksha Desai, Snehit Prabhu, Eric Yang, Michael G. Ozawa, Paul J. Hanson, Fang Wu, Matthew J. Frank, John H. Baird, Lori Muffly, Gursharan K. Claire, Juliana Craig, Katherine A. Kong, Dhananjay Wagh, John Coller, Sylvia K. Plevritis, Bita Sahaf, David B. Miklos, Crystal L. Mackall. Reverse fate mapping of CD19-targeted CAR T cells in patients with large B-cell lymphoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3603.