Background
Adoptive transfer of T cell receptor gene-engineered T (TCR-T) cells can induce durable anti-cancer responses. TBI-1301 is a novel gene therapy produced by engineering autologous lymphocytes to express an NY-ESO-1-specific TCR using a retrovirus vector encoding siRNA to silence endogenous TCR. In this study, we characterize long-lived persisting transferred TBI-1301 TCR-T cells following adoptive transfer at the single-cell level.
Methods
Patients eligible for the approved study (UHN REB 15-9534) included those with informed consent, HLA-A*02:01 or A*02:06 haplotype, and NY-ESO-1 expression by IHC. PBMCs were harvested and processed to generate engineered TBI-1301 TCR-T cells. Patients received an infusion of 5×109 cells on day 0 after lymphodepletion with cyclophosphamide (CY) alone (750 mg/m2 on day -7 and -6) or in combination with fludarabine (FLU) (30 mg/m2 on day -7 and -6). Endpoints included safety, efficacy, and biological correlates for persistence of TCR-T cells post-infusion. The TBI-1301 infusion product and persisting TBI-1301 TCR-T cells were assessed by multi-parameter flow cytometry, single-cell RNA and TCR sequencing analysis.
Results
Clinical activity of TBI-1301 has previously been described. In the CY only cohort, 5/9 experienced grade 1–2 CRS, and, for the CY/FLU cohort, 3/5 patients experienced grade 2 CRS. The RECIST response rate in synovial sarcoma patients, whose tumors express high levels of NY-ESO-1, was 28.6%. In patients receiving CY alone, persistence beyond 100 days was detected in 3 patients at low levels (0.03–0.05% of CD8 T cells). In contrast, higher levels of persistence (5.6–7.7% of CD8 T cells) were observed in 2 patients receiving CY/FLU. Long-lived TBI-1301 cells exhibited a naïve/memory phenotype expressing CD45RA and CCR7. Single cell RNA sequencing analysis enabled the tracking of the infused TBI-1301 T cells from day 0 to day 125, including clonotypic cells sharing the same TCR sequences. UMAP visualization revealed that persisting TCR-T are distinct from infused cells, with higher expression of TCF7, LEF1, and CCR7 in both peripheral CD8+ and CD4+ TCR-T cells. In CD8+ T cells on day 125, differentiation trajectories of endogenous and TCR-T cells were recapitulated using potential of heat diffusion for affinity-based transition embedding (PHATE) and Slingshot pseudotime. Single-cell gene signature scoring revealed that TCR-T cells were enriched with a Tstem gene signature, whereas endogenous T cells exhibited a Tpex gene signature.
Conclusions
These data demonstrate sustained persistence of infused TBI-1301 T cells. Understanding the cell states of persisting TCR-T cells provides valuable insights for developing combination approaches to enhance the efficacy of new therapeutics.
Trial Registration
ClinicalTrials. gov NCT02869217.
Ethics Approval
Subjects underwent informed consent to a University Health Network Research Ethics Board protocol, UHN REB # 15-9534.