Observational Study Of PNH Clone Size By High Sensitivity Flow Cytometry In High-Risk Patients InA Large Russian Population
Authors: Babenko Elena , Sipol Alexandra , Borisov Vyacheslav , Naumova Elena , Boyakova Elena , Yakunin Dimitry , Glazanova Tatyana , Chubukina Zhanna , Pronkina Natalya , Popov Alexander , Saveliev Leonid , Partylova Elena , Vlasova Larisa , Mustafin Ilshat , Ramazanov Bulat , Galtseva Irina , Fidarova Zalina , Lugovskaya Svetlana , Lisukov Igor , Kulagin Alexander 
Published: October 2019

Introduction

Paroxysmal nocturnal hemoglobinuria (PNH) is a rare and life-threatening hematopoietic stem cell disease caused by a partial or absolute deficiency of proteins linked to the cell surface membrane via a glycosylphosphatidyl-inositol anchor, which leads to complement-induced intravascular hemolysis mediated via the membrane attack complex. Multiparameter high-sensitivity flow cytometric measurement of PNH clones is the method of choice for the diagnosis of PNH, as recommended by the International Clinical Cytometry Society (ICCS). After publication of the ICCS guidelines, screening of patients considered at high risk of PNH was commenced in Russia. Data are presented on PNH clone size distribution across patients with relevant ICD-10 diagnostic codes (based on patients′ initial assumed diagnoses).

Methods

Patients were tested for the presence and size of PNH clones using high-sensitivity flow cytometry across nine laboratories. PNH clone evaluations were performed as described in the ICCS guidelines: CD59/CD235a monoclonal antibodies for RBC; CD45/CD15/CD24/FLAER for granulocytes and; CD45/CD64/CD14/FLAER or CD45/CD33/CD14/FLAER for monocytes. The sensitivity for PNH clone detection was 0.01%. Changes in PNH clone size were evaluated among patients who had follow-up studies after initial measurements.

Results

1889 patients were assessed between October 2011 and June 2013 (Table 1). Suspected PNH and bone-marrow disorders (AA, MDS, cytopenia) were the most common reasons for PNH testing. The greatest proportions of patients with PNH clones were among those with of an initial assumed diagnosis of AA or PNH. Notably, around 40% of patients with an initial assumed diagnosis of PNH actually had no detectable PNH clones. Most patients with small clone sizes (< 1%) were in the AA, MDS and hemolytic anemia groups. Overall, mean clone sizes were slightly higher in monocytes (31.5%) than in granulocytes (30.1%) across the diagnostic categories. While there was generally a good correlation between clone size measurements in granulocytes and monocytes (linear regression r2 = 0.9851), 10% of PNH-positive patients had detectable clones only in one of these leucocyte populations (i.e. either in monocytes or in granulocytes, but not both). PNH clones in RBCs were generally lower than in granulocytes. Repeat clone size measurements were performed in 316 patients over a mean follow-up period of 7.8 months. In patients with initial clone sizes <50% the PNH clones tended to decrease over time, whereas in patients with initial clone sizes >50%, clones tended to increase. PNH clones were not changed at all in 98 patients at follow-up, among whom 48% were patients with AA.

Conclusion

These screening data confirm the utility of high-sensitivity flow cytometry testing in high-risk patient groups to ensure early and accurate diagnosis and to aid in the effective clinical management of patients.

Disclosures:

Babenko: Alexion: Research Funding. Sipol:Alexion: Research Funding. Borisov:Alexion: Employment. Naumova:Alexion: Research Funding. Boyakova:Alexion: Research Funding. Glazanova:Alexion: Research Funding. Chubukina:Alexion: Research Funding. Pronkina:Alexion: Research Funding. Popov:Alexion: Research Funding. Mustafin:Alexion: Research Funding. Fidarova:Alexion: Honoraria. Lisukov:Alexion: Honoraria. Kulagin:Alexion: Honoraria.