Multicolor Flow Cytometry and Multigene Next-Generation Sequencing Are Complementary and Highly Predictive for Relapse in Acute Myeloid Leukemia after Allogeneic Transplantation.
Minimal residual disease (MRD) in acute myeloid leukemia (AML) is typically measured using multiparameter flow cytometry (MFC). Detection of leukemia mutations using multigene next-generation sequencing (NGS) can potentially be used to measure residual disease. We used a targeted 28-gene NGS panel to detect mutations and different-from-normal 10-color MFC to measure MRD in AML patients before allogeneic hematopoietic stem cell transplantation (HCT). Residual disease was defined when any abnormal blast population was detected using MFC and when any leukemia allele was detected with a variant allele frequency (VAF)  ≥ 5% using NGS. We tracked the clearance of leukemia alleles between AML diagnosis and immediately before HCT and found that mutations in DNMT3A, TET2, and JAK2 were less likely to be cleared than NPM1, IDH 1/2, and FLT3-ITD. Despite varying sensitivities, the concordance rate of residual disease detection before HCT using the 2 assays was 44 of 62 (71%) evaluable cases. Discordance could be explained by residual mutations in DNMT3A and TET2 that were not detected by MFC and presence of residual leukemia mutations with VAF below the established thresholds for mutation calling. Presence of flow MRD and residual mutations immediately before HCT using the 2 assays was associated with relapse risk (MFC: hazard ratio,  4.62; 95% confidence interval [CI], 1.32 to 16.09; P = .016 and NGS: hazard ratio,  4.35; 95% CI, 1.63 to 11.6; P = .003) and survival (MFC: hazard ratio,  2.44; 95% CI, 1 to 5.97; P = .05 and NGS: hazard ratio, 2.1; 95% CI, .97 to 4.55; P = .059) after HCT. Residual disease detected concurrently by MFC and NGS conferred the highest relapse risk compared with patients who were either negative by both assays or had discordant status (overall, P = .008). Although MFC is universally applicable, a multigene NGS approach to measuring residual disease in AML provides additional information on differential clearance of disease alleles and can assess clonal architecture before transplantation.
Bone Marrow Transplant Service, Memorial Sloan Kettering Cancer Center, New York, New York. Electronic address: gettab@mskcc.org.
Bartlomiej M
BM
Getta
Department of Epidemiology and Biostatistics, Memorial Sloan Kettering Cancer Center, New York, New York.
Sean M
SM
Devlin
Leukemia Service, Memorial Sloan Kettering Cancer Center, New York, New York; Memorial Sloan Kettering Center for Hematologic Malignancies, New York, New York.
Ross L
RL
Levine
Diagnostic Molecular Pathology Laboratory, Memorial Sloan Kettering Cancer Center, New York, New York.
Maria E
ME
Arcila
Diagnostic Molecular Pathology Laboratory, Memorial Sloan Kettering Cancer Center, New York, New York.
Abhinita S
AS
Mohanty
Diagnostic Molecular Pathology Laboratory, Memorial Sloan Kettering Cancer Center, New York, New York.
Ahmet
A
Zehir
Leukemia Service, Memorial Sloan Kettering Cancer Center, New York, New York.
Martin S
MS
Tallman
Bone Marrow Transplant Service, Memorial Sloan Kettering Cancer Center, New York, New York.
Sergio A
SA
Giralt
Department of Laboratory Medicine, Memorial Sloan Kettering Cancer Center, New York, New York.
Mikhail
M
Roshal
D000328:Adult; D000368:Aged; D005260:Female; D005434:Flow Cytometry; D018380:Hematopoietic Stem Cell Transplantation / Q000379:methods*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000175:diagnosis*; D008297:Male; D008875:Middle Aged; D018365:Neoplasm, Residual / Q000175:diagnosis*; D000090243:Nucleophosmin; D012008:Recurrence; D019172:Transplantation Conditioning / Q000379:methods*; D014184:Transplantation, Homologous / Q000379:methods*; D055815:Young Adult
Acute myeloid leukemia; Flow cytometry; Minimal residual disease; Next-generation sequencing