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Myeloperoxidase in myelodysplastic syndromes
sis. Fifty imputed data sets were created with a total run length of 50,000 iterations and imputations made every 1,000 iterations.
We quantified the accuracy of each neutrophil MPO expression parameter in discriminating MDS and non- MDS patients by estimating the area under the ROC curve. We compared the area under the ROC curve for each parameter with that for the RCV. The significance probability was adjusted for multiple comparisons using the Bonferroni method.
The specificity, positive and negative predictive values, and likelihood ratios of the test results were estimated across a range of RCV values that achieved sensitivity of from 100% to 90% in the retrospective case control study. Since neutrophil MPO expression in PB would be mainly used to rule out MDS, we selected a threshold with a like- lihood ratio for a negative test result point estimate that was lower than 0.10.28
Two-tailed P<0.05 was considered statistically signifi- cant. Analyses were performed using Stata Special Edition version 14.0 (Stata Corporation, College Station, TX, USA).
Results
Retrospective case control study
Forty-four MDS patients and 44 controls were included in the study. The mean age for all patients was 73.3 years (standard deviation, 10.4), and 38 (43%) were female (Table 1). MDS with excess blasts, MDS with multilineage dysplasia, and CMML accounted for 55% (24 of 44), 20% (9 of 44), and 11% (5 of 44) of all MDS patients, respec- tively (Table 2). MDS cases had lower median hemoglobin concentration, platelet counts, and absolute neutrophil counts than controls (Table 1).
Compared with controls, MDS cases yielded compara- ble median and mean values, but a higher RCV for neu-
trophil MPO expression measured by flow cytometric analysis in peripheral blood (Table 1). Odds ratios of MDS associated with a 1% increase in RCV were 1.80 (95%CI: 1.39-2.33) in univariable analysis and 2.22 (95%CI: 1.31- 3.76) in multivariable analysis adjusting for age, C-reactive protein, and creatinine concentrations. RCV values for neutrophil MPO expression in PB were elevated across all WHO classification MDS types, ranging from 28.3% (in a patient with MDS with multilineage dysplasia) to 99.3% (in a patient with MDS with isolated del(5q)) (Table 2). Median RCV values for MPO expression of circulating neutrophils were 41.1% [interquartile range (IQR): 38.6- 47.2] and 38.6% (IQR: 36.6-46.0) for 25 low- and 19 high- risk MDS patients, compared with 30.9% (IQR: 29.7-31.9) for 44 controls (Online Supplementary Table S1).
The area under the ROC curve (0.94, 95%CI: 0.86-0.97) for the RCV was higher than that for median and mean (Figure 3). These findings were unchanged after excluding CMML cases (Online Supplementary Table S2). Sensitivity point estimates ranged from 100% to 91% for RCV thresholds varying between 28% and 32% (Table 3). A RCV value <30% yielded a negative predictive value of 93% and a likelihood ratio of a negative test result of 0.07 (Table 3). All cases but one with established MDS diagno- sis had RCV values >30%. The exception was a 72-year old female case with multilineage dysplasia, for whom isolated peripheral thrombocytopenia (94x109/L) and a 28.3% RCV value for MPO expression in the PB neu- trophil population were found. RCV value <28.0%, there- fore, excluded MDS with both sensitivity and negative predictive value estimates of 100%, but occurred in a small proportion of patients (3.4%, 3 of 88).
Prospective validation study
Sixty-eight consecutive patients referred for suspected MDS were included in the validation cohort study. The mean age for all patients was 74.7 years (standard devia- tion, 9.2), and 29 (43%) were female (Table 4). The preva-
AB
Figure 2. Monoparametric histograms of peripheral blood neutrophil myeloperoxidase (MPO) expression. Values are: mean, fluorescence intensity (FI); median, FI; and robust coefficient of variation (RCV), %. (A) Control subject. (B) Myelodysplastic syndrome case.
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