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Novel Duplex Sequencing Assay Improves Detection of MRD in AML

By Patrick Daly - Last Updated: March 21, 2024

The novel DuplexSeq™ AML-XP, an error-corrected, next-generation sequencing assay, was “highly sensitive, specific, accurate, and precise” for detecting measurable residual disease (MRD) in acute myeloid leukemia (AML), according to an analytical validation led by Mark McElwain, PhD, of TwinStrand Biosciences in Seattle, Washington.

Though 2022 European LeukemiaNet guidelines define positive MRD status as ≥0.1% variant allele frequency (VAF), “recent evidence suggests that identification of lower frequency variants may be informative for managing subsequent therapy,” Dr. McElwain and colleagues stated.

The assay detected variants down to 0.00055% VAF with a limit of detection of 0.0098% for single-nucleotide variants and insertions and deletions and a limit of blank of 0%, the researchers reported in a presentation at the 2024 Tandem Transplantation & Cellular Therapy Meetings of ASTCT® and CIBMTR®.

The validation utilized human genomic DNA carrying 25 targeted coding-sequence variants as well as DNA from AML-positive blood and bone marrow samples. The authors wrote that 82% of targeting regions showed sequencing depth more than 80% of the panel-wide mean, and more than 99.5% of targeted regions showed depth more than 20% of the mean.

In the limit of detection analyses for variants with more than 0.0098% VAF, the assay had overall percent agreement of 97.8%, positive percent agreement of 98.3%, and negative percent agreement of 97.7%. Results were linear going from 0.0016% to 96.1% VAF (R2=0.95). Additionally, all six expected FLT3-ITD variants and five expected NPM1 insertions were detected.

“The assay was highly repeatable and reproducible across two operators, two lots of reagents, and three independent runs of library preparation,” according to the presentation.

In closing, “study of recurrence rates and outcomes in patients with less than 0.1% VAF using [duplex sequencing] has the potential to improve outcomes through earlier detection of residual disease,” Dr. McElwain suggested.


McElwain M, Olson JM, Nguyen K, et al. An AML targeted duplex sequencing assay that can detect measurable residual disease (MRD) at a sensitivity better than 0.01% variant allele frequency. Abstract #122. Presented at the 2024 Tandem Transplantation & Cellular Therapy Meetings of ASTCT® and CIBMTR®; February 21-24, 2024; San Antonio, Texas.

Original Source: Novel Duplex Sequencing Assay Improves Detection of MRD in AML | Blood Cancers Today

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