Quality Control of Flow Cytometry Data Analysis for Evaluation of Minimal Residual Disease in Bone Marrow From Acute Leukemia Patients During Treatment

Abstract

Low levels of leukemia cells in the bone marrow, minimal residual disease (MRD), are considered to be a powerful indicator of treatment response in acute lymphatic leukemia (ALL). A Nordic quality assurance program, aimed on standardization of the flow cytometry MRD analysis, has been established before implementation of MRD at cutoff level 10 as one of stratifying parameters in next Nordic Society of Pediatric Hematology and Oncology (NOPHO) treatment program for ALL. In 4 quality control (QC) rounds 15 laboratories determined the MRD levels in 48 follow-up samples from 12 ALL patients treated according to NOPHO 2000. Analysis procedures were standardized. For each QC round a compact disc containing data in list-mode files was sent out and results were submitted to a central laboratory. At cutoff level 10, which will be applied for clinical decisions, laboratories obtained a high concordance (91.6%). If cutoff level 10 was applied, the concordance would be lower (85.3%). The continuing standardization resulted in better concordance in QC3 and QC4 compared with QC1 and QC2. The concordance was higher in precursor B as compared with T-cell ALL. We conclude that after standardization, flow cytometry MRD detection can be reliably applied in international, multicenter treatment protocols.

Prospective comparison of two flow cytometry methodologies for monitoring minimal residual disease in a multicenter treatment protocol of childhood acute lymphoblastic leukemia

“…Modulation of several markers is another confounding variable important for MRD analysis as we have seen in some of our discordant cases and as was reported before (30). Interestingly, similar to the NOPHO group (29) we observed that the concordance was higher in precursor B when compared with T-cell ALL.…”

“…Per time point best qualitative agreement was in exchanged LMD from D-15 (86%) as opposed to those from D-33 (52%). Also in the NOPHO QC study (29) at a cutoff level 10, laboratories obtained a high concordance (91.6%). Remarkably, in our study, with no prior standardization and utilizing two very different FCM-MRD methodologies, qualitative agreement was 80% for D-33 and quantitative agreement of D-15 samples was 80.8%.…”

Detection of minimal residual disease in pediatric acute lymphoblastic leukemia

“…Despite differences in cytometers and software usage, blinded inter-laboratory tests of list-mode data interpretation gave highly concordant results (intraclass correla- robustness of this approach in refining MRD-based stratification with measurements on day 15 (37). The feasibility of standardized multicentric MRD assessment has also been shown by others groups (66,67). Six-eightcolor-FCM MRD techniques are currently employed for leukemia immunophenotyping.…”

Standardization of Flow Cytometric Minimal Residual Disease Monitoring in Children With B-Cell Precursor Acute Lymphoblastic Leukemia. Russia–belarus Multicenter Group Experience

“…Существен-ными недостатками мониторинга МОБ методом про-точной цитометрии по сравнению с количественной полимеразной цепной реакцией (ПЦР) являются субъективность и сложность стандартизации в рамках многоцентровых исследований [8,14]. В то время как технология молекулярно-генетического ис следования МОБ хорошо отработана и стандартизована [8,14,15], опубликовано лишь несколько работ по начальным этапам стандартизации цитометрического мониторин-га ОЛЛ в рамках отдельных исследовательских групп [16][17][18].…”

“…Однако сложности применения данного варианта ПЦР определяют необходимость разработ-ки стандартизованных протоколов определения МОБ проточной цитометрией в рамках различных терапев-тических протоколов. На данный момент такие по-пытки проводятся группами AIEOP-BFM [16], UKALL [17], NOPHO [18]. Кроме того, высокий уровень стан-дартизации определения МОБ при ОЛЛ предлагает не связанная с определенными терапевтическими протоколами группа EuroFlow [14].…”