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Single-Cell DNA Sequencing and Immunophenotypic Profiling to Track Clonal Evolution in an Acute Myeloid Leukemia Patient

María García-Álvarez, Ana Yeguas, Cristina Jiménez, Alejandro Medina-Herrera, Verónica González-Calle, Montserrat Hernández-Ruano, Rebeca Maldonado, Irene Aires, Cristina Casquero, Inmaculada Sánchez-Villares, Ana Balanzategui, María Eugenia Sarasquete, Miguel Alcoceba, María Belén Vidriales, Marcos González-Díaz, Ramón García-Sanz, María Carmen Chillón
Biomedicines Dec 2023

Single-cell DNA sequencing can address the sequence of somatic genetic events during myeloid transformation in relapsed acute myeloid leukemia (AML). We present an NPM1-mutated AML patient with an initial low ratio of FLT3-ITD (low-risk ELN-2017), treated with midostaurin combined with standard chemotherapy as front-line treatment, and with salvage therapy plus gilteritinib following allogenic stem cell transplantation after relapse. Simultaneous single-cell DNA sequencing and cell-surface immunophenotyping was used in diagnostic and relapse samples to understand the clinical scenario of this patient and to reconstruct the clonal composition of both tumors. Four independent clones were present before treatment: DNMT3A/DNMT3A/NPM1 (63.9%), DNMT3A/DNMT3A (13.9%), DNMT3A/DNMT3A/NPM1/FLT3 (13.8%), as well as a wild-type clone (8.3%), but only the minor clone with FLT3-ITD survived and expanded after therapy, being the most represented one (58.6%) at relapse. FLT3-ITD was subclonal and was found only in the myeloid blast population (CD38/CD117/CD123). Our study shows the usefulness of this approach to reveal the clonal architecture of the leukemia and the identification of small subclones at diagnosis and relapse that may explain how the neoplastic cells can escape from the activity of different treatments in a stepwise process that impedes the disease cure despite different stages of complete remission.




Institution Type


Indication / Modality

Acute Myleoid Leukemia, Acute Myleoid Leukemia (AML)

Goal of Study

Clinical Profiling, CHIP, Therapeutic Resistance

PAD Project


Analytes Assessed

Extracellular Protein, InDels, SNV

Sample Prep

Whole Cells

Proof Point Demonstrated

Clonality, Co-occurrence, Multi-omics