Contributions
Abstract: EP725
Type: E-Poster Presentation
Session title: Gene therapy, cellular immunotherapy and vaccination - Biology & Translational Research
Background
Pivotal trials of Axicabtagene-Ciloleucel (Axi-cel) and Tisagenlecleucel (Tisa-cel) for the treatment of relapsed and refractory (r/r) Diffuse Large B-Cell Lymphoma (DLBCL) demonstrated complete responses in 40%>58% of the patients with accompanied grade ≥3 adverse events in approximately 30% of the patients. Recently, others could associate high tumor volumne and low CAR T-cell expansion in vivo with poor treatment outcome. We hypothesize, that expansion of CAR T-cells and T cells in vivo depend on the composition and differentiation status of (CAR) T-cell subsets and that differential expansion of (CAR) T cell subsets determines response rate.
Aims
We performed a retrospective, single-center study in which we characterized potential determinants of CAR T-cell and T-cell expansion[MS1] and their association with treatment response at time of transfusion and 28 days thereafter: i) the distribution of CD4+ and CD8+ (CAR) T-cells subsets ii) the memory phenotype and immune checkpoint expression of (CAR) T-cells.
Methods
Patients with r/r DLBCL were treated with the CD19 specific CAR T-cell products Axi-cel or Tisa-cel at the LMU. Peripheral blood from 38 patients was collected before, during and after CAR T transfusion. Multi-parameter flow cytometry was done to detect CAR T-cells and assess marker for T-cell differentiation (CD3, CD4, CD8, CD45RA, CCR7) and immune checkpoints (PD-1, TIM-3, LAG-3). CAR T-cells were quantified based on a two step-staining approach with a biotinylated recombinant CD19 protein. Efficacy was evaluated through PET-CT scan three months and by CT scan six months after CAR T-cell transfusion and patients were categorized in Responder (complete remission) and Non-Responder (partial remission, stable or progressive disease).
Results
CAR T-cell expansion peaked between day 7 and 14 in the peripheral blood with a significantly higher expansion of CD8+ CAR T-cells compared to CD4+ CAR T-cells between day 4 and 28 (p=0.004-0.007). CAR T-cells were detectable in most patients until day 105 (sensitivity 1:1000). Responder (R) displayed a higher absolute number of CD8+ CAR T-cells over time after transfusion compared to Non-Responder (NR, p=0.02). CAR T-cells in R showed a higher frequency of an effector (p=0.004) and lower frequency of a central memory phenotype (p=0.002) compared to NR 28 days after CAR T-cell transfusion. Furthermore, assessment of immune checkpoints over time revealed a lower expression of PD-1 (p<0.0001), TIM-3 (p<0.0001), LAG-3 (p=0.001) on CAR T-cells in R compared to NR.
Conclusion
We have successfully established an Immunmonitoring platform for the assessment of CAR T-cell expansion and their immune phenotypes in vivo through flow cytometry. We observed relationships of CAR T-cell kinetics, memory subsets and immune checkpoint expression with efficacy. These findings might bear the potential to monitor treatment responses and prevent disease relapse together with other clinical variables.
Keyword(s): CAR-T, Diffuse large B cell lymphoma, Flow cytometry, T cell expansion
Abstract: EP725
Type: E-Poster Presentation
Session title: Gene therapy, cellular immunotherapy and vaccination - Biology & Translational Research
Background
Pivotal trials of Axicabtagene-Ciloleucel (Axi-cel) and Tisagenlecleucel (Tisa-cel) for the treatment of relapsed and refractory (r/r) Diffuse Large B-Cell Lymphoma (DLBCL) demonstrated complete responses in 40%>58% of the patients with accompanied grade ≥3 adverse events in approximately 30% of the patients. Recently, others could associate high tumor volumne and low CAR T-cell expansion in vivo with poor treatment outcome. We hypothesize, that expansion of CAR T-cells and T cells in vivo depend on the composition and differentiation status of (CAR) T-cell subsets and that differential expansion of (CAR) T cell subsets determines response rate.
Aims
We performed a retrospective, single-center study in which we characterized potential determinants of CAR T-cell and T-cell expansion[MS1] and their association with treatment response at time of transfusion and 28 days thereafter: i) the distribution of CD4+ and CD8+ (CAR) T-cells subsets ii) the memory phenotype and immune checkpoint expression of (CAR) T-cells.
Methods
Patients with r/r DLBCL were treated with the CD19 specific CAR T-cell products Axi-cel or Tisa-cel at the LMU. Peripheral blood from 38 patients was collected before, during and after CAR T transfusion. Multi-parameter flow cytometry was done to detect CAR T-cells and assess marker for T-cell differentiation (CD3, CD4, CD8, CD45RA, CCR7) and immune checkpoints (PD-1, TIM-3, LAG-3). CAR T-cells were quantified based on a two step-staining approach with a biotinylated recombinant CD19 protein. Efficacy was evaluated through PET-CT scan three months and by CT scan six months after CAR T-cell transfusion and patients were categorized in Responder (complete remission) and Non-Responder (partial remission, stable or progressive disease).
Results
CAR T-cell expansion peaked between day 7 and 14 in the peripheral blood with a significantly higher expansion of CD8+ CAR T-cells compared to CD4+ CAR T-cells between day 4 and 28 (p=0.004-0.007). CAR T-cells were detectable in most patients until day 105 (sensitivity 1:1000). Responder (R) displayed a higher absolute number of CD8+ CAR T-cells over time after transfusion compared to Non-Responder (NR, p=0.02). CAR T-cells in R showed a higher frequency of an effector (p=0.004) and lower frequency of a central memory phenotype (p=0.002) compared to NR 28 days after CAR T-cell transfusion. Furthermore, assessment of immune checkpoints over time revealed a lower expression of PD-1 (p<0.0001), TIM-3 (p<0.0001), LAG-3 (p=0.001) on CAR T-cells in R compared to NR.
Conclusion
We have successfully established an Immunmonitoring platform for the assessment of CAR T-cell expansion and their immune phenotypes in vivo through flow cytometry. We observed relationships of CAR T-cell kinetics, memory subsets and immune checkpoint expression with efficacy. These findings might bear the potential to monitor treatment responses and prevent disease relapse together with other clinical variables.
Keyword(s): CAR-T, Diffuse large B cell lymphoma, Flow cytometry, T cell expansion