Clinical Reflection of Immunophenotyping in Paroxysmal Nocturnal Hemoglobinuria: Experience of a Single Center
Authors: Sahin Ugur , Merter Mustafa , Ataca Pinar , Atilla Erden , Atesagaoglu Berna , Pekcan Gultekin , Civriz Bozdag Sinem , Toprak Selami Kocak , Topcuoglu Pervin , Arslan Onder , Ozcan Muhit , Demirer Taner , Ilhan Osman , Akan Hamdi , Beksac Meral , Konuk Nahide , Gurman Gunhan 
Published: October 2019

Introduction:

Flow cytometric assays (FCA), especially FLAER based methods, have become important recently in means of diagnosis of paroxysmal nocturnal hemoglobinuria (PNH). In this retrospective study, we aimed to describe diagnostic features of PNH patients along with disease characteristics.

Methods:

FCA results of patients ordered for PNH diagnosis between November 2007 and July 2014 were retrospectively evaluated. FCA results were expressed as percentage of granulocytes and monocytes negative for CD55/59. The FLAER/CD24-CD14 assay has been available in our center since December 2011 and used for confirmation of FCA results since then. The distribution of erythrocyte types according to CD59 expression (type 1, 2 and 3) were also given in percentages.

Results:

FCA for PNH diagnosis was performed for a total of 175 patients. FLAER method was available in 136 of these assays (77.7%). A PNH clone was not detected in 136 (77.7%) of the FCAs. PNH clone free patients were diagnosed with unclassified anemia (n=34, 19.4%), other unclassified cytopenias (n=30, 17.2%), non-PNH hemolytic anemias (n=7, 4.0%), myelodysplastic syndrome (n=23, 13.1%), aplastic anemia (n=9, 5.1%), primary myelofibrosis (n=1, 0.6%), leukemia (n=7, 4.0%), lymphoma (n=3, 1.7%), thrombotic incidents (n=14, 8.0%), rheumatological disorders (n=4, 2.3%) and unknown diagnosis (n=4, 2.3%).

A PNH clone was detected in 39 patients (22.3%). Of these, 27 (69.2%) constituted classical PNH and 12 (30.8%) PNH accompanying other bone marrow disorders. In the PNH clone positive group, the accompanying bone marrow disorders were aplastic anemia (n=9) and myelodysplastic syndrome (n=3). Median age at diagnosis was 31 (10-82) for classical PNH and 35 (18-77) for PNH accompanying other bone marrow disorders.

Erythrocyte transfusion requirement was prominent in 48.7% (n=19) of the patients and thrombotic incidents occured in 20.5% (n=8). Leukemic transformation did not take place in any of the patients. In one patient with aplastic anemia, the FCA became positive for PNH clone within one year of follow-up.

Among classical PNH patients, thrombosis was observed only among patients with a clone size of > 50%. Clone size (both for granulocyte and monocyte clones) correlated with corrected reticulocyte and serum lactate dehydrogenase levels (for granulocytes p=0.001 and p<0.001, for monocytes p=0.001 and p<0.001, respectively).

Treatment modalities of the PNH clone positive patients included eculizumab (n=12, 30.8%), allogeneic stem cell transplantation (n=13, 33.3%) and none/other (CsA, steroids, ATG, danazol) (n=16, 41.0%).

Conclusion:

FCA is an important modality for the diagnosis and follow-up of PNH patients. Clone size might be used to assess the severity of the disease and the risk of thrombosis in classical PNH. In patients with PNH accompanying other bone marrow disorders clone sizes are generally small and complications seen in classical PNH are rare. However, aplastic anemia and myelodysplastic syndrome might evolve into PNH during the disease course, which is reflected as an increase in clone size. Thus, FCA might be used in selected patients with aplastic anemia and myelodysplastic syndrome, especially when treatment failure or unexpected complications are suspected.

Disclosures

No relevant conflicts of interest to declare.