Measurable Residual Disease (MRD) refers to a group of cancer cells that persist after treatment and is a crucial harbinger of disease relapse in acute myeloid leukemia and other malignancies. Understanding the biological characteristics that enable MRD clones to resist treatment is essential for guiding the development of more effective therapies. Distinguishing residual leukemia clones, pre-leukemic clones, and normal precursors remains a challenge for current MRD tools. In this study, the team developed a single-cell MRD (scMRD) detection method by combining flow cytometry enrichment of targeted precursor/blast populations with integrated single-cell DNA sequencing and immunophenotyping. Our scMRD detection demonstrated high sensitivity, approximately 0.01%, enhancing MRD detection while elucidating the clonal architecture of surviving hematopoietic/pre-leukemic cells and leukemia cells.
Why is MRD of interest?
Quantifying Measurable Residual Disease (MRD) provides vital prognostic information for acute myeloid leukemia (AML). There is a wide array of platforms for MRD detection, each varying in sensitivity and suitability for individual patients. MRD serves as a biomarker for AML prognosis as it can predict rates of treatment response, survival, and relapse. Using MRD as an alternative endpoint in clinical trial design can significantly expedite drug approvals.
The Challenges of NGS in MRD Testing
While these testing methods are highly valuable, there is still a pressing clinical need to distinguish residual leukemia cells from mutated CH/pre-leukemic cells, which do not always predict relapse. Residual subclones may possess different leukemia potential, while ancestral clones may only result in CH without overt AML, a distinction that cannot be accurately described through bulk MRD analysis. Additionally, extensive NGS MRD testing can detect mutations in mature populations that lack leukemia potential, which is common in patients undergoing differentiation-inducing therapies. These biological complexities necessitate careful interpretation of MRD analysis, ultimately hindering the clinical application of bulk sequencing and flow cytometry-based methods.
The Advantages of This New Technology
The research team has developed a novel multi-target single-cell MRD (scMRD) detection method. This method combines flow cytometry enrichment of targeted precursor/blast populations with integrated single-cell DNA sequencing and immunophenotyping. Furthermore, they have developed a computational algorithm based on single-nucleotide polymorphisms (SNPs) to deconvolve multiplexed data from parallel samples.
In simple terms, the process begins by using Fluorescence-Activated Cell Sorting (FACS) to enrich live CD34+ and/or CD117+ progenitor cells (including CD34+CD117-, CD34+CD117+, and CD34-CD117+). Subsequently, these cells are loaded onto the Mission Bio Tapestri single-cell sequencing platform.
This study demonstrates the feasibility of enumerating and characterizing MRD using single-cell genotype and immunophenotype profiling through progenitor cell enrichment and scDNA+ protein technology. scMRD detection not only enhances the sensitivity of MRD detection but also achieves sufficient resolution to characterize the clonal architecture of leukemia precursor/leukemia cells persisting after treatment, potentially increasing the specificity of MRD detection. This technology provides a more reliable screening method for AML prognosis and treatment.
