Phenotype and functional state of endogenous T-cells support T-cell engager therapy in the post-CAR T therapy setting Free

Several new therapies have been approved for the treatment of B-cell NHL in the relapsed/refractory setting. CD3xCD20 bispecific antibodies and chimeric antigen receptor T-cell (CAR T) therapy are powerful immunotherapies that activate T-cells to kill CD20- or CD19-expressing malignant cells. Approved CAR T therapies rely on structural engineering and adoptive transfer of autologous T-cells following lymphodepletion, while CD3xCD20 bispecifics are engineered antibodies that recruit and redirect the activity of endogenous T-cells against tumor. The aim of this study is to describe the phenotypic composition of endogenous T-cells and CAR T post-CAR T therapy and assess function of these cells ex vivo in response to CD3xCD20 bispecifics.

Clinical response data, endogenous T-cell characteristics, and ex vivo activity of viably frozen peripheral blood mononuclear cells (PBMCs) in the presence of CD3xCD20 bispecific antibodies were analyzed from 22 patients (diffuse large B-cell lymphoma [DLBCL], n=19; follicular lymphoma [FL], n=3) from the Montpellier University Hospital after written informed consent in accordance with the Declaration of Helsinki and institutional research board approval. Patients received axicabtagene ciloleucel (Yescarta, Gilead Sciences; n=17) or tisagenlecleucel (Kymriah, Novartis; n=5), both CD19-targeted CAR T. In this study, all patients received CAR T therapy after ≥2 prior lines of therapy and CAR T was detected using flow cytometry from the Montpellier University Hospital. In ex vivo experiments, both glofitamab and epcoritamab were tested for methods development and feasibility. Epcoritamab was then used to assess functionality of the full cohort of PBMC samples collected five days before (D -5; at the time of apheresis), and at 3 to 6 months (M3, M6) after CAR T therapy. Phenotypic assessments were performed throughout therapy by flow cytometry including D -5, after the standard conditioning regimen containing cyclophosphamide and fludarabine 2 to 3 days based on the CAR T therapy, and up to 300+ days post-CAR T therapy.

In this CAR T-treated cohort, complete responses were observed in 81.8% (n=18/22) with durability of complete response from 7 months to 4 years and included patients with DLBCL and FL. Four patients relapsed within 3 to 6 months with only 1 of the early relapsed patients showing no CAR T expansion. Endogenous T-cells and CAR T were independently assessed and were found to expand with different kinetics, peak at 7 to 10 days for CAR T, and later (~10–300+ days) for endogenous T-cells. In paired analysis pre- to post-CAR T infusion, CD8 T-cells expanded and CD4 T-cells decreased in both responders and nonresponders. In patients with complete response, a significant expansion of endogenous CD8+ effector memory T-cells (P<.01) was observed without increase in LAG3, PD-1, or TIM3. A similar trend was observed in patients with progressive disease. Endogenous T-cells in PBMC samples collected pre- (D -5) and post- (M3, M6) CAR T therapy showed similar ex vivo cytotoxic activity and exhibited similar induction of the activation antigen CD25 in response to either epcoritamab, glofitamab, or anti-CD3/CD28 in both CAR T responders and nonresponders.

Endogenous T-cells post-CAR T therapy in DLBCL and FL were functionally responsive to epcoritamab. Phenotypic changes post-CAR T therapy were consistent with T-cell reconstitution following lymphopenia post-cytoreduction where the T-cells proliferate in response to homeostatic cytokines that favor CD8+ T-cells with a specific effector/effector memory phenotype (Williams et al. Semin Immunol, 2007). The phenotypic changes were independent of the kinetics and response to CAR T therapy, suggesting that bispecific engagement can activate and redirect T-cells even in patients who failed CAR T therapy. This data supports the benefit of epcoritamab post-CAR T therapy, as evidenced by the high complete response rate observed in epcoritamab-treated CAR T-exposed patients (Thieblemont C, et al. Leukemia 2024).