Baseline Immune State and T Cell Clonal Kinetics Are Associated with Response to CAR-T Therapy in Large B-Cell Lymphoma

Engineered adoptive cellular therapy with CD19-targeting chimeric antigen receptor T-cells (CART) has revolutionized the treatment landscape for patients with relapsed/refractory (R/R) large B cell lymphoma (LBCL); however, failure rates remain high. Thus, improved molecular understanding of patient and treatment characteristics linked to response are urgently needed to define the basis for achieving sustained remission. Single cell technologies have the power to evaluate heterogenous populations and provide unprecedented ability to track T cell clonal kinetics. Here we focused on patients with R/R LBCL treated with axicabtagene ciloleucel (axi-cel), from whom we collected serial peripheral blood mononuclear cell (PBMC) samples from Day -30 to +28 relative to CAR-T infusion, as well as bag washings from infusion products (IPs). We evaluated 308,357 single cell transcriptomes (10x Genomics) from serial PBMCs and IPs from 29 patients (16 responders [R], 13 non-responders [NR], R defined as complete [CR] or partial response [PR] at 1 year, NR defined as progressive disease [PD] by 1 year), of which 220,653 originated from 19 patients in a previously reported cohort generated from our center (Haradhvala et al., Nat Med, 2022) and 87,704 newly generated from 10 of 50 patients (29 R, 21 NR) from an extension cohort on whom flow cytometry and mass cytometry (CyTOF) were also generated. With this expanded power, we systematically evaluated baseline and immediate post-treatment samples for features associated with durable response. While we observed no quantitative differences in post-infusion circulating immune subsets in R versus NR, scRNA-seq analysis yielded two notable findings at baseline (Day -30). First, the proportion of B cells (mostly naïve B cells) was increased in R (n=12) compared to NR (n=7) (p=0.012). Protein expression by flow cytometry in the full extension cohort (of whom 20 [13 R, 7 NR] had available PBMC data at baseline) revealed a trend towards increased B cells in R (p=0.16), dominated by four exceptional outliers in whom B cells comprised 2-30% of total PBMCs, all of whom were CR. Resegregating these 20 patients to 11 CR vs 9 non-CR confirmed higher frequency of circulating B cells (median CR 0.18% vs non-CR 0.011%, p=0.0057). Patients with B cells had longer time from anti-CD20 therapy (60, 41, 22, and 10 months) whereas patients without B cells had median 1 month since last anti-CD20 therapy. Second, lymphocyte to monocyte ratio [ALC/AMC]