Biomarkers of Response to Anti-CD19 Chimeric Antigen Receptor (CAR) T-Cell Therapy in Patients with Chronic Lymphocytic Leukemia

Abstract

The adoptive transfer of autologous T cells genetically modified to express a CD19-specific, 4-1BB/CD3z-signaling CAR (CTL019) has shown remarkable activity and induce long-term remissions in a subset of patients with relapsed/refractory chronic lymphocytic leukemia (CLL). To date, little is known about predictive indicators of efficacy. This study was designed to evaluate biomarkers of clinical response to CTL019 in CLL. We studied forty-one patients with advanced, heavily pre-treated and high-risk CLL who received at least one dose of CTL019 cells. We show that in vivo expansion and persistence are key quality attributes of CTL019 cells in CLL patients who have complete responses to therapy; in 2 patients responses are sustained beyond five years and accompanied by the persistence of functional CTL019 cells. Furthermore, durable remissions were associated with transcriptomic signatures of early memory T cells, while T cells from non-responding patients were enriched in genes belonging to known pathways of terminal differentiation and exhaustion. Polychromatic flow cytometry also demonstrated a significantly higher level of T cell exhaustion markers on the infused CAR T cells and reduced CD27 expression in non-responding patients. Accordingly, the combined assessment of PD1 and CD27 expression on CD8+ CTL019 cells in the infusion product accurately predicted response to treatment. Restimulation of the infusion product through the CAR further demonstrated that CTL019 cells from complete responders secreted significantly higher levels of several cytokines, including CCL20, IL-21, IL-22, IL-17, and IL-6, suggesting that the STAT3 signaling pathway may play a role in potentiating the enhanced potency of CTL019 cells. To identify a phenotype of T cells that is predictive of response prior to CTL019 manufacturing, we initially retrospectively evaluated the proportions of naïve, stem cell memory, central memory, effector memory and effector cells at the time of leukapheresis and observed either marginally significant or no significant correlations with clinical outcome. A systematic, unbiased analysis of the same biomarker panel revealed that the frequency of CD27+CD45RO- cells in the CD8+ T cell population correlated significantly with complete and durable responses to this therapy. Analysis of the infusion products using the same flow cytometric panel showed that most (>95%) of T cells expressed CD45RO at the end of the manufacturing run; CD27 expression frequencies, however, were maintained at the same level as in the leukapheresis. Together, these findings suggest that intrinsic T cell fitness dictates both response and resistance to highly active engineered CAR T cells. Thus, enrichment of T cells with optimal differentiation potential and proliferative capacity by timing of collection or culture modification might potentiate the generation of maximally efficacious infusion products. These data and additional immunological biomarkers may be used to identify which patients are most likely to respond to adoptive transfer strategies, leading to an enhanced personalized approach to cellular therapy. Disclosures Fraietta: Novartis: Patents & Royalties: Novartis, Research Funding. Lacey:Novartis: Research Funding. Wilcox:Novartis: Research Funding. Bedoya:Novartis: Research Funding. Chen:Novartis: Research Funding. Orlando:Novartis: Employment. Brogdon:Novartis: Employment. Hwang:Novartis: Research Funding. Frey:Novartis: Research Funding; Amgen: Consultancy. Pequignot:Novartis: Research Funding. Ambrose:Novartis: Research Funding. Levine:Novartis: Patents & Royalties, Research Funding; GE Healthcare Bio-Sciences: Consultancy. Bitter:Novartis: Employment. Porter:Genentech: Employment; Novartis: Patents & Royalties, Research Funding. Xu:Novartis: Research Funding. June:Immune Design: Consultancy, Equity Ownership; University of Pennsylvania: Patents & Royalties; Celldex: Consultancy, Equity Ownership; Novartis: Honoraria, Patents & Royalties: Immunology, Research Funding; Tmunity: Equity Ownership, Other: Founder, stockholder ; Pfizer: Honoraria; Johnson & Johnson: Research Funding. Melenhorst:Novartis: Patents & Royalties: Novartis, Research Funding.

A novel antibody-TCR (AbTCR) platform combines Fab-based antigen recognition with gamma/delta-TCR signaling to facilitate T-cell cytotoxicity with low cytokine release

“…S5 ). Retrospective analysis from published CAR-T clinical studies have found that T cells that are more naïve, less differentiated, and less exhausted correlate with improved efficacy 17 , 18 . Interestingly, after CD3/CD28 expansion, but before antigen engagement, a greater fraction of ET190L1-AbTCR-T cells displayed a naive (T N ) and stem cell memory (T SCM ) T cell phenotype compared to ET190L1-CAR-T cells (Figs.…”

Cell therapy products: focus on issues with manufacturing and quality control of chimeric antigen receptor T‐cell therapies

“…Immunological dogma and phenotypic and transcriptomic profiling data support the thesis that less differentiated central memory (CD45RO+, CD62L+, CD95+) and pluripotent stem cell (CD45RA+, CD62L+, CD95+) memory T‐cell subsets may be optimal in this regard . Fraietta et al . investigated biomarkers in 41 Chronic Lymphoid Leukaemia (CLL) patients treated with Kymriah.…”

“…116,117 Immunological dogma and phenotypic and transcriptomic profiling data support the thesis that less differentiated central memory (CD45RO+, CD62L+, CD95+) and pluripotent stem cell (CD45RA+, CD62L+, CD95+) memory T-cell subsets may be optimal in this regard. [73][74][75] Fraietta et al 101 investigated biomarkers in 41 Chronic Lymphoid Leukaemia (CLL) patients treated with Kymriah. Durable remissions were associated with transcriptomic signatures of early memory T-cells, while T-cells from non-responding patients were enriched in genes belonging to known pathways of terminal differentiation and exhaustion.…”

The Pharmacology of T Cell Therapies

“…Recent retrospective analysis of CTL019 data suggest that immunophenotypic characterization of T cell composition of both the starting leukopheresis material as well as the infused T cell product are predictive of engraftment and response. 30 In addition, efforts to generate T cell products with a more defined composition have shown good engraftment results. 31 However, because they have also not been directly compared to products with less defined composition, it is difficult to know how much benefit is derived from these approaches.…”