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

Myelodysplastic Syndromes Clinical Trial

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

Status Active, not recruiting

Clinical Trial 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.

Clinical Trial Description

The primary objective of MPO-MDS-develop study is to estimate the discriminative accuracy (i.e., area under the ROC curve along with 95% CI) of a standardized and semi-automatic flow cytometric assay of neutrophil MPO expression in peripheral blood for the diagnosis of MDS in adult patients.

The secondary objectives are to assess:

1. intra-laboratory reproducibility (intra- and inter-assay precision) for intra-individual RCV measurement of neutrophil MPO expression in peripheral blood;

2. specimen stability for intra-individual RCV measurement of neutrophil MPO expression in peripheral blood at different time points in comparison with H0 (stored at 4°C and at room temperature);

3. inter-laboratory reproducibility for intra-individual RCV measurement of neutrophil MPO expression in peripheral blood;

4. the negative predictive value for intra-individual RCV measurement of neutrophil MPO expression in peripheral blood (with a prespecified threshold of 30.0%) in ruling out MDS in consecutive adult patients.

The MPO-MDS-Develop project is a multicenter, cross-sectional, diagnostic accuracy study of an index test by comparison with a reference standard, in unselected consecutive patients.

Screening: All consecutive patients referred to the immuno-hematology lab at the study sites for suspicion of MDS will be screened for eligibility. A lab physician will review inclusion and exclusion criteria, using computerized medical and laboratory records.

Recruitment: Participants will be included in the study once all the screening activities have been conducted and only if the patient meets all inclusion and none exclusion criteria. The consent for flow cytometry analysis of peripheral blood sample and data collection through chart review will be sought under a regime of "non-opposition" (opt-out): after appropriate written information is delivered, cross-sectional data will be collected except in case of opposition from the patient. All patients included in the study will be assigned a unique patient identification number. This number will be used to identify the patient throughout the study.

Index test: Flow cytometry analysis of neutrophil myeloperoxidase expression in peripheral blood will be performed using a single ready-to-use tube with lyophilized antibodies developed in partnership with Becton Dickinson France SAS and blinded to the reference standard.

Reference standard: The diagnosis of MDS will be established according to the World Health Organization (WHO) classification, based on clinical data, peripheral blood cytopenia, cytomorphology of peripheral blood and bone marrow aspirate, and cytogenetic analysis. The criteria for MDS diagnosis are 1) the presence of ≥10% dysplastic cells in any hematopoietic lineage, 2) the exclusion of acute myeloid leukemia (defined by the presence of ≥20% peripheral blood or bone marrow blasts), and 3) the exclusion of reactive etiologies of dysplasia. Cytopenia is defined by hemoglobin concentration <10 g/dL, platelet count <100x109/L, and/or absolute neutrophil count <1.8 x109/L. Yet a diagnosis of MDS could be made with milder levels of cytopenia. Idiopathic cytopenia of uncertain significance (ICUS) is defined by unexplained mild persistent cytopenia for at least 6 months and the failure to establish the diagnosis to MDS according to published guidelines. Consistent with WHO classification, MDS subcategorization will rely on the degree of dysplasia (unilineage versus multilineage), blast percentages, presence of ring sideroblasts, and cytogenetic analysis (del(5q)). The criteria for CMML diagnosis are 1) the presence of persistent peripheral blood monocytosis ≥1 x109/L, and 2) monocyte accounting for more than 10% of the white blood cell differential count. Evaluation of bone marrow cytomorphology will be performed prospectively by experienced hematopathologists who are blinded to the index test results.

Patients with confirmed suspicion of MDS: Participants for whom the diagnosis of MDS (or CMML) is confirmed by the reference standard will be categorized as patients with confirmed suspicion of MDS.

Patients with unconfirmed suspicion of MDS: Participants for whom the diagnosis of MDS (or CMML) is ruled out by the reference standard will be categorized as patients with unconfirmed suspicion of MDS. This latter subgroup will include patients with ICUS, as defined in accordance with published guidelines.

Follow-up: No follow-up visit is planned in this cross-sectional diagnostic accuracy study. ;

Related Conditions & MeSH terms

• Myelodysplastic Syndromes
• Preleukemia
• Syndrome

• NCT number: NCT04399018
• Study type: Observational
• Source: University Hospital, Grenoble
• Status: Active, not recruiting
• Start date: July 27, 2020
• Completion date: December 2024