Flow Cytometric MRD Assessment in Acute Lymphoblastic Leukemias

Acute lymphoblastic leukemia (ALL) is one of the very first malignancy where the assessment of early response to therapy by minimal/measurable residual disease (MRD) monitoring has proven to be cardinal tool for guiding therapeutic choices. At present, MRD detection is not only used for the assessment of initial treatment response and subsequent risk stratification but also for monitoring disease burden in the setting of hematopoietic stem cell transplant. Multicolor flow cytometry (FCM) for the assessment of MRD has been in existence for more than two decades. It is presently the most commonly used technique worldwide for MRD assessment in ALL. The technique has evolved from two to three color assays in its early phases to eight and more color assays in present time, which enables detection of one leukemic cell in 10 boolean AND 4 or more cells. The assessment of MRD is based on analysis of expression of lineage-associated markers and either looking at "leukemia associated immunophenotypes" or identify "different from normal" patterns. A rapid turn-around-time and direct quantification of viable residual leukemic cells are advantages of FCM over molecular techniques of MRD assessment. On the other hand, one of the prime limitations of detection of residual cells by FCM is the immunophenotypic shifts that are observed as a result of chemotherapeutic reagents. In addition, introduction of immunotherapy, especially against important gating markers like CD19, has posed significant challenge to FCM-based MRD assays, and requires modification of antibody panels for an alternate gating and analysis strategy. Finally, standardization and validation of MRD assay and use of internal and external quality controls are extremely important aspects for a clinical laboratory providing MRD reports for patient care.