Monitoring of kinetics and exhaustion markers of circulating CAR-T cells as early predictive factors in patients with B-cell malignancies
Clara García-Calderón, Belén Sierro-Martínez, Estefanía García‐Guerrero, Luzalba Sanoja-Flores, Raquel Muñoz-García, Victoria Ruiz-Maldonado, María Reyes Jiménez‐León, Javier Delgado‐Serrano, Águeda Molinos-Quintana, Beatriz Guijarro-Albaladejo, Inmaculada Carrasco-Brocal, José-Manuel Lucena, José-Raúl Garcı́a-Lozano, Cristina Blázquez-Goñi, Juan Luis Reguera, Marı́a-Francisca González-Escribano, Marta Reinoso‐Segura, Javier Briones, José Antonio Pérez‐Simón, Teresa Caballero‐Velázquez
Abstract
Purpose: CAR-T cell therapy has proven to be a disruptive treatment in the hematology field, however, less than 50% of patients maintain long-term response and early predictors of outcome are still inconsistently defined. Here, we aimed to optimize the detection of CD19 CAR-T cells in blood and to identify phenotypic features as early biomarkers associated with toxicity and outcomes. Experimental design: In this study, monitoring by flow cytometry and digital PCR (dPCR), and immunophenotypic characterization of circulating CAR-T cells from 48 patients treated with Tisa-cel or Axi-cel was performed. Results: Validation of the flow cytometry reagent for the detection of CAR-T cells in blood revealed CD19 protein conjugated with streptavidin as the optimal detection method. Kinetics of CAR-T cell expansion in blood confirmed median day of peak expansion at seven days post-infusion by both flow cytometry and digital PCR. Circulating CAR-T cells showed an activated, proliferative, and exhausted phenotype at the time of peak expansion. Patients with increased expansion showed more severe CRS and ICANs. Immunophenotypic characterization of CAR-T cells at the peak expansion identified the increased expression of co-inhibitory molecules PD1 and LAG3 and reduced levels of the cytotoxicity marker CD107a as predictors of a better long-term disease control. Conclusions: These data show the importance of CAR-T cells in vivo monitoring and identify the expression of PD1LAG3 and CD107a as early biomarkers of long-term disease control after CAR-T cell therapy.