A complementary role of multiparameter ﬂow cytometry and high-throughput sequencing for minimal residual disease detection in chronic lymphocytic leukemia: an European Research Initiative on CLL study

Leukemia, 2016 · DOI: 10.1038/leu.2015.313 · Published: January 29, 2016

Cardiovascular Science Oncology Research Methodology & Design

Simple Explanation

This study aimed to improve the detection of minimal residual disease (MRD) in chronic lymphocytic leukemia (CLL) using flow cytometry and high-throughput sequencing. A standardized flow cytometry assay was developed that can reliably detect CLL cells at very low levels (0.0010%). High-throughput sequencing showed good agreement with flow cytometry and could detect even lower levels of CLL cells (1 in a million).

Study Duration

Not specified

Participants

128 samples from 108 patients with CLL or monoclonal B-cell lymphocytosis

Evidence Level

Not specified

Key Findings

1
A six-marker flow cytometry panel (CD19, CD20, CD5, CD43, CD79b, and CD81) was identified as reliable and convenient for identifying typical CLL cells.
2
The developed flow cytometry assay can reliably detect residual CLL cells down to 0.0010% (10 −5) with a single-tube assay.
3
High-throughput sequencing (HTS) using the ClonoSEQ assay showed good concordance with flow cytometry at the 0.010% (10 −4) level and good linearity to a detection limit of 1 in a million (10 −6).

Research Summary

This study by the European Research Initiative on CLL (ERIC) identified and validated a flow-cytometric approach to reliably quantitate CLL cells to the level of 0.0010% (10 −5). A parallel analysis of high-throughput sequencing using the ClonoSEQ assay showed good concordance with ﬂow cytometry results at the 0.010% (10 −4) level, and also provides good linearity to a detection limit of 1 in a million (10 −6). The combination of both technologies would permit a highly sensitive approach to MRD detection while providing a reproducible and broadly accessible method to quantify residual disease and optimize treatment in CLL.

Practical Implications

Improved MRD Detection

The study provides a more sensitive and reliable method for detecting minimal residual disease in CLL patients, potentially leading to better risk stratification and treatment decisions.

Treatment Optimization

Using MRD as a surrogate endpoint could expedite the evaluation of new CLL therapies and help optimize treatment strategies aimed at disease eradication.

Wider Accessibility

The single-tube flow cytometry assay, with its platform-independent reagent specification, can be easily adopted by more laboratories, improving access to MRD testing for CLL patients.

Study Limitations

1
The development of this assay was primarily based on peripheral blood analysis because of the logistical difﬁculty in obtaining normal bone marrow.
2
The variation in quantiﬁcation between ﬂow cytometry and HTS could be higher than preferable and thus further work is warranted to standardize the quantitative analysis of HTS.
3
CLL-associated IGH sequences are frequently unmutated and stereotyped (i.e. identical sequences in different patients) and may be present in the normal IG repertoire.

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