Single-cell profiling identifies pre-existing CD19-negative subclones in a B-ALL patient with CD19-negative relapse after CAR-T therapy

Tracy Rabilloud,Marie Loosveld,Delphine Potier,Mathis Nozais,Dominique Payet-Bornet,Saran Pankaew

doi:10.1038/s41467-021-21168-6

Abstract

Chimeric antigen receptor T cell (CAR-T) targeting the CD19 antigen represents an innovative therapeutic approach to improve the outcome of relapsed or refractory B-cell acute lymphoblastic leukemia (B-ALL). Yet, despite a high initial remission rate, CAR-T therapy ultimately fails for some patients. Notably, around half of relapsing patients develop CD19 negative (CD19neg) B-ALL allowing leukemic cells to evade CD19-targeted therapy. Herein, we investigate leukemic cells of a relapsing B-ALL patient, at two-time points: before (T1) and after (T2) anti-CD19 CAR-T treatment. We show that at T2, the B-ALL relapse is CD19 negative due to the expression of a non-functional CD19 transcript retaining intron 2. Then, using single-cell RNA sequencing (scRNAseq) approach, we demonstrate that CD19neg leukemic cells were present before CAR-T cell therapy and thus that the relapse results from the selection of these rare CD19neg B-ALL clones. In conclusion, our study shows that scRNAseq profiling can reveal pre-existing CD19neg subclones, raising the possibility to assess the risk of targeted therapy failure.

Highlights

B-cell acute lymphoblastic leukemia (B-ALL) is a hematologic malignancy arising from uncontrolled proliferation of B lymphoblasts
The B-ALL patient underwent a relapse after chemotherapy and CD19pos leukemic cells from bone marrow (BM) were harvested (T1 cells = before chimeric antigen receptors (CARs)-T cell therapy; Supplementary Fig. 1a)
After an initial complete remission, the patient experienced a frank relapse, as noted by circulating blasts and an infiltration (>90%) of CD10posCD19neg leukemic cells in the BM (Supplementary Fig. 1b) from which cells were harvested (T2 cells = after CAR-T cell therapy). We asked whether these CD19neg B-ALL clones were present before CAR-T cell therapy

Summary

Introduction

B-cell acute lymphoblastic leukemia (B-ALL) is a hematologic malignancy arising from uncontrolled proliferation of B lymphoblasts. Regarding the treatment of relapsed/refractory B-ALL, adoptive T cells expressing a CAR directed against the B-cell-associated surface marker, CD19, have been used in several clinical trials[8,9,10,11,12]. Those studies showed that CAR-T therapies achieved high remission rates ranging from 70 to 90% at shortterm. The CD19pos relapse mainly results from the low potency and poor persistence of CAR-T cells[13] To overcome these issues, efforts are made to optimize clinical strategies and CAR-T cells engineering[14,15]. Our work reveals the presence of preexisting CD19neg B-ALL clones before the CAR-T treatment

Methods

Results

Conclusion