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Publications

Monitoring of Leukemia Clones in B-cell Acute Lymphoblastic Leukemia at Diagnosis and During Treatment by Single-cell DNA Amplicon Sequencing


Meyers S, Alberti-Servera L, Gielen O, Erard M, Swings T, De Bie J, Michaux L, Dewaele B, Boeckx N, Uyttebroeck A, De Keersmaecker K, Maertens J, Segers H, Cools J, Demeyer S.
HemaSphere Mar 2022
Abstract

Acute lymphoblastic leukemia (ALL) is characterized by the presence of chromosomal changes, including numerical changes, translocations, and deletions, which are often associated with additional single-nucleotide mutations. In this study, we used single cell-targeted DNA sequencing to evaluate the clonal heterogeneity of B-ALL at diagnosis and during chemotherapy treatment. We designed a custom DNA amplicon library targeting mutational hotspot regions (in 110 genes) present in ALL, and we measured the presence of mutations and small insertions/deletions (indels) in bone marrow or blood samples from 12 B-ALL patients, with a median of 7973 cells per sample. Nine of the 12 cases showed at least 1 subclonal mutation, of which cases with PAX5 alterations or high hyperdiploidy (with intermediate to good prognosis) showed a high number of subclones (1 to 7) at diagnosis, defined by a variety of mutations in the JAK/STAT, RAS, or FLT3 signaling pathways. Cases with RAS pathway mutations had multiple mutations in FLT3, NRAS, KRAS, or BRAF in various clones. For those cases where we detected multiple mutational clones at diagnosis, we also studied blood samples during the first weeks of chemotherapy treatment. The leukemia clones disappeared during treatment with various kinetics, and few cells with mutations were easily detectable, even at low frequency (<0.1%). Our data illustrate that about half of the B-ALL cases show >2 subclones at diagnosis and that even very rare mutant cells can be detected at diagnosis or during treatment by single cell-targeted DNA sequencing.

VIEW

Area

Heme

Institution Type

Academia

Indication / Modality

Acute Lymphoblastic Leukemia (ALL)

Goal of Study

Clonal Evolution, Longitudinal, Retrospective, MRD, Clonal Heterogeneity, Therapeutic Resistance

Key Genes

NRAS, KRAS, BRAF, PTPN11, FLT3, JAK1, CREBBP, SETD2, EZH2, KMT2D, IKZF1, PAX5, OVGP1

PAD Project

No

Analytes Assessed

InDels, SNV

Sample Storage

Fresh Frozen

Sample Prep

Whole Cells

Sample Type

PBMC, Patient Material

Tissue / Organ

Bone Marrow Aspirates

Species

Human

Panel Used

ALL Published Panel

Proof Point Demonstrated

Better than Bulk, Clonality, Co-occurrence, Phylogeny, Sensitivity
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