Clonal expansion of CAR T cells harboring lentivector integration in the CBL gene following anti-CD22 CAR T-cell therapy

Abstract

1. Reexpansion of CAR T cells led to further investigations which confirmed the clonal nature of this expansion.

Graphical abstract

Figure 1.

Clinical course. (A) FDG–positron emission tomography (PET) scan demonstrating significant extramedullary disease burden at pretreatment, followed by clinical response at day +30, and subsequent full resolution of any PET-avid disease at day +90. Subject was MRD⁻ in the bone marrow by the day +30 evaluation and remained MRD⁻ at the subsequent time points. (B) Clinical timeline with relevant laboratory findings. During the initial CRS, patient had a concurrent rise in CRP and ferritin which coincided with the initial WBC peak. During the second expansion phase, the patient remained clinically well without any fevers and had only modest increase in C-reactive protein (CRP) and ferritin, but with an even higher WBC than with the initial expansion. Concurrent table demonstrates the % of T cells which were CAR⁺ in the peripheral blood at the various timepoints.

Figure 2.

Expansion and persistence of CAR T cell with lentivector integrated in the CBL gene. (A) TCR diversity of CAR product before infusion; samples taken from the patient on days +51 and +53. A major clonotype that was obvious in day +51 and became more dominant on day +53. (B) Integration site diversity of CAR product before infusion and samples taken from the patient on days +51 and +53. The top clones at days +51 and +53 are the same and have same vector integrated in chr11 at position 119143577. (C) Details of the integration in the dominant clone at days +51 and +53. The lentiviral vector carrying the EF1a–anti-CD22 CAR integrated in the second intron of CBL gene on chr11, in the opposite orientation to the gene. (D) Longitudinal quantification of CAR vector copy numbers and the CBL clone in patient WBCs by ddPCR. The vector copy number (copies per 100 cells) shows 2 phases of expansion of CAR T cells. The first phase peaked at day +7 to +14 and subsided around day +30. The second phase peaked at day +53. The clone with the vector inserted into the CBL gene was quantified with ddPCR assay specific for the junction between the vector and host DNA. The clone is detectable, at a low level, from day +7 throughout day +53. During the second phase of CAR T expansion, the CBL clone became the dominant clone at day +53, constituting 46% of total WBCs in the patient and more than one-half of the CAR T cells.