Clonal origin of KMT2A wild-type lineage-switch leukemia following CAR-T cell and blinatumomab therapy

Abstract

Children with acute lymphoblastic leukemia (ALL) undergoing anti-CD19 therapy occasionally develop acute myeloid leukemia (AML). The clonal origin of such lineage-switch leukemias^1-4 remains unresolved. Here, we reconstructed the phylogeny of multiple leukemias in a girl who, following multiply relapsed ALL, received anti-CD19 cellular and antibody treatment and subsequently developed AML. Whole genome sequencing unambiguously revealed the AML derived from the initial ALL, with distinct driver mutations that were detectable before emergence. Extensive prior diversification and subsequent clonal selection underpins this fatal lineage switch. Genomic monitoring of primary leukemias and recurrences may predict therapy resistance, especially regarding anti-CD19 treatment.

Fig. 1. The clonal relationship between AML and ALL.

a, Schematic overview of different relations between an ALL and subsequent AML from the same patient and corresponding mutational readouts. (i) The AML and ALL may be clonally unrelated malignancies, sharing no mutations, as is the case for treatment-related AML (t-AML). (ii) They may arise from a common precursor cell, typified by shared somatic mutations likely including a first driver mutation, but is itself not a cancer cell. (iii) The AML may derive from the ALL, which has undergone a true lineage switch through transdifferentiation, typified by the AML harboring all somatic mutations of the primary ALL. Note that asynchronous presentations of ALL and AML may involve mutations in the second lesion induced by the therapy for the first lesion. b, A fish plot showing the clone size of the different ALL and AML clones over the course of the clinical history of the patient. Number in parentheses after sample identifier refers to the genome-wide coverage of sequencing. mths, months; MDS, myelodysplastic syndrome. c, Reconstructed phylogeny of clones with annotated putative driver mutations and CNVs. Number of single base substitution mutations underlying each branch of the phylogenetic tree are denoted above the branch. Dashed lines group clones into initial ALL, ALL relapses and AML relapses. fs, frameshift; del, deletion; rearr., rearrangement.

Fig. 2. Mutational patterns across leukemia samples.

a, Copy-number state in the PAX5-ZCCHC7 locus, revealing a single loss in the initial ALL and a subsequent second hit in the relapse samples with nearby, but unique break points resulting in a constitutive loss of PAX5 and ZCCHC7. Both deletions were flanked by recombination signal sequence (RSS) motifs, indicating that these deletions were likely RAG-mediated. b, Exposures of the three identified signatures in this patient, with signature A corresponding to COSMIC reference SBS1, signature B to SBS5 and signature C to SBS87. The latter is associated with thiopurine exposure. Only branches with 100 SNVs or more are displayed. c, Schematic of treatment courses of the patient, along with thiopurine-related mutagenesis. As the progression root and the ALL relapse root (its descendant) harbor thiopurine-induced mutations, the split between the ALL and AML lineages must have happened during the treatment for the initial ALL. d, Estimated clone size of ancestral AML lineage across samples, as estimated from interrogating all mutant sites in diploid regions (n = 2,318) in the AML root branch (Methods). The red line indicates the background error rate estimated from 32 unmatched blood samples. Error bars denote the 95% CI around maximum likelihood binomial estimate of clone size. ***P < 0.001, one-sided binomial test with null hypothesis that number of variant counts is drawn from background error distribution. The exact P value for the ALL R2 sample is 1.5 × 10⁻¹⁵¹ and 0 for all AML-related samples. e, Genome-wide copy-number profile of the two AML samples, obtained 3 months apart, showing that the later AML acquired extensive aneuploidy in a short time, likely facilitated by the biallelic loss of TP53.

Extended Data Fig. 1. Heatmap of VAFs.

Heatmap of the variant allele frequency of all identified substitutions across all samples. Branch colors correspond to Fig. 1b–c.

Extended Data Fig. 2. VAF plots in replicate samples.

Plot of the variant allele frequency of mutations between different samples from the same time point: initial ALL (a) and All relapse 1 (b). Mutations are coloured by their branch assignment.

Extended Data Fig. 3. Mutational signatures.

Profiles of the three identified mutational signatures in this patient, with Signature A corresponding to COSMIC reference SBS1, Signature B to SBS5 and Signature C to SBS87. The latter is associated with thiopurine exposure.