Development and Prospective Validation of a Standardized Flow Cytometric Assay of Peripheral Blood Neutrophil Myeloperoxidase Expression for Ruling Out Myelodysplastic Syndromes.

Summary

Myelodysplastic syndromes (MDS) comprise a heterogeneous group of clonal bone marrow neoplasms that predominate in the elderly, with a median age at diagnosis of 70 years. The diagnosis of MDS relies on peripheral blood cytopenia and morphologic dysplasia for one or more hematopoietic cell lineage. Cytopenia is evidenced with hemogram while dysplasia requires bone marrow aspirate, which is an invasive procedure .

Considering the low prevalence of disease among subjects referred for suspected MDS, many patients are exposed to unnecessary bone marrow aspiration-related discomfort and harms. Therefore, an objective assay based on a peripheral blood sample that accurately discriminates MDS from other cytopenia etiologies is highly desirable.

We have previously developed and refined a flow cytometric analysis protocol for quantifying neutrophil MPO expression in peripheral blood at three university-affiliated hospitals (i.e., Clermont-Ferrand, Saint-Etienne, and Grenoble) (Raskovalova et al, Hematologica 2019). We found that the robust coefficient of variation (RCV, computed as the robust standard deviation divided by the median) within an individual subject was the best parameter in discriminating patients with versus without MDS.

Although promising, flow cytometric analysis of neutrophil MPO expression in peripheral blood is technically complex, time consuming, and not standardized. Hence, its performance requires specific expertise and the results show substantial variability. A single ready-to-use tube with lyophilized antibodies would have the potential to standardize the measurement of neutrophil MPO expression in peripheral blood across laboratories, with results available within 30-60 min in routine practice.

In this study, the investigators hypothesize that a standardized and semi-automatic flow cytometric assay of neutrophil MPO expression in peripheral blood could accurately rule out MDS and obviate the need for bone marrow aspiration and biopsy, with sensitivity and negative predictive value estimates approaching 100%.

In this observational diagnostic accuracy study, burden will be null for recruited patients. No specific intervention is assigned to participants. All diagnostic testing, procedures, and medication ordering are performed at the discretion of attending physicians. A test result will have no impact on patient management. .Compliance with current guidelines disseminated by the French Haute Autorité de Santé (HAS) will be advocated for the diagnostic work-up of patients with suspected MDS. No follow-up visits are planned in this cross-sectional study.

Conditions

• Myelodysplastic Syndromes

Interventions

OTHER

Diagnostic Test: Flow cytometry analysis of neutrophil myeloperoxidase expression

Flow cytometry analysis of neutrophil myeloperoxidase expression in peripheral blood samples will be performed within 24 h of MDS diagnostic evaluation and blinded to the reference standard. Peripheral blood samples will be collected in 5 ml (EDTA) anticoagulant plastic tubes and processed within 24 h maximum of collection. Blood sample will be stained according to the manufacturers' recommendations with a lyophilized cocktail ("LyotubeTM ready-to-use", BD Bioscience). At least 10,000 neutrophils will be acquired on a 3-laser, 8-color BD FACSCanto-II TM flow cytometer (BD Biosciences, San José, CA). Each marker will be expressed as median, geometric and arithmetic mean, regular and robust coefficient of variation.

Sponsors & Collaborators

• University Hospital, Grenoble

  lead OTHER

Principal Investigators

• Tatiana Raskovalova, MD · Centre Hospitalier Universitaire Grenoble Alpes

• Richard Veyrat-Masson, MD · Centre Hospitalier Universitaire Clermont Ferrand

• Carmen Aanei, MD · Centre Hospitalier Universitaire Saint-Étienne

Eligibility

Min Age

18 Years

Max Age

100 Years

Sex

ALL

Healthy Volunteers

No

Timeline & Regulatory

Start

2020-07-27

Primary Completion

2024-08-31

Completion

2024-12-31

Countries

• France

Study Locations