Contributions
Abstract: EP712
Type: E-Poster Presentation
Session title: Enzymopathies, membranopathies and other anemias
Background
PKD is the second most frequent enzymopathy and the first to produce chronic hemolytic anemia. Its prevalence is underestimated due to the low clinical suspicion, especially in mild cases, the low expressiveness of the peripheral blood smear and the limitations of enzymatic techniques. Next-generation sequencing techniques (NGS) allow simultaneous study of multiple causal genes in multiple patients, which provides the diagnosis of congenital hemolytic anemias.
Aims
During one year, all cases of undiagnosed chronic hemolytic anemias suspected to be PKD in Spain were remitted for NGS analysis.
Methods
DNA obtained from peripheral leukocytes was analyzed using a 40 genes panel covering congenital anemias at the Hematology Services of the Clínico San Carlos and Cruces University Hospitals. Ilumina platform (Truseq Custom Amplicon) was used. Pathogenic variants and those of uncertain significance were reported, using the GRCh37 / Hg19 genome as reference.
Results
Ninety-nine patients with congenital anemias from 23 national centers were analyzed. PKD mutation was detected in 24 patients, 18 of them correctly suspected of PKD (18/24, 75%), 3 of them were associated with a membranopathy, while 3 other suspected cases (12.5%) were finally diagnosed as hereditary membrane disorders, particularly xerocytosis (HX, 2 cases). Initial suspicion was different from PKD in 75 patients (18 membranopathies, 4 G6PD, 4 CDA and 49 with unknown origin). PKD explained 4% of the cases of undiagnosed anemias (2/49) and was also detected in two patients suspected of membranopathy (2/18), interestingly with these two different diseases associated. It should be noted that half (2/4) of the suspected cases of congenital dyserythropoietic anemia (CDA) were eventually diagnosed as PKD. No cases of PKD were detected in patients suspected of G6PD deficiency.Twenty-eight different gene variants were found in the 24 PKD confirmed cases. Six variants were constantly repeated in more than one patient, sometimes in combination: R486W (50% of patients), R337Q (17%), E241X (12.5%), R510Q (8%), I378X (8%), R559X (8%).
Conclusion
Our results confirms that NGS technique has become a powerful diagnostic tool, enabling a diagnoses in 78% of patients with congenital anemias in this study. Further, it allowed for the diagnoses of the majority of patients with clinical suspicion of PKD (75%), the detection of combined genetic alterations (in particular, 5 out of 24 PKD cases were combined with membranopathies), and the diagnosis of PKD in a high percentage of cases suspected of CDA (50%) or in a low percentage of unexplained anemias (4%). NGS also enabled the correct identification of two XH cases, initially suspected of PKD. The pathogenic variants detected in Spain are consistent with those previously reported, with the exception of the lower percentage of R510Q.
Keyword(s): Pyruvate kinase deficiency
Abstract: EP712
Type: E-Poster Presentation
Session title: Enzymopathies, membranopathies and other anemias
Background
PKD is the second most frequent enzymopathy and the first to produce chronic hemolytic anemia. Its prevalence is underestimated due to the low clinical suspicion, especially in mild cases, the low expressiveness of the peripheral blood smear and the limitations of enzymatic techniques. Next-generation sequencing techniques (NGS) allow simultaneous study of multiple causal genes in multiple patients, which provides the diagnosis of congenital hemolytic anemias.
Aims
During one year, all cases of undiagnosed chronic hemolytic anemias suspected to be PKD in Spain were remitted for NGS analysis.
Methods
DNA obtained from peripheral leukocytes was analyzed using a 40 genes panel covering congenital anemias at the Hematology Services of the Clínico San Carlos and Cruces University Hospitals. Ilumina platform (Truseq Custom Amplicon) was used. Pathogenic variants and those of uncertain significance were reported, using the GRCh37 / Hg19 genome as reference.
Results
Ninety-nine patients with congenital anemias from 23 national centers were analyzed. PKD mutation was detected in 24 patients, 18 of them correctly suspected of PKD (18/24, 75%), 3 of them were associated with a membranopathy, while 3 other suspected cases (12.5%) were finally diagnosed as hereditary membrane disorders, particularly xerocytosis (HX, 2 cases). Initial suspicion was different from PKD in 75 patients (18 membranopathies, 4 G6PD, 4 CDA and 49 with unknown origin). PKD explained 4% of the cases of undiagnosed anemias (2/49) and was also detected in two patients suspected of membranopathy (2/18), interestingly with these two different diseases associated. It should be noted that half (2/4) of the suspected cases of congenital dyserythropoietic anemia (CDA) were eventually diagnosed as PKD. No cases of PKD were detected in patients suspected of G6PD deficiency.Twenty-eight different gene variants were found in the 24 PKD confirmed cases. Six variants were constantly repeated in more than one patient, sometimes in combination: R486W (50% of patients), R337Q (17%), E241X (12.5%), R510Q (8%), I378X (8%), R559X (8%).
Conclusion
Our results confirms that NGS technique has become a powerful diagnostic tool, enabling a diagnoses in 78% of patients with congenital anemias in this study. Further, it allowed for the diagnoses of the majority of patients with clinical suspicion of PKD (75%), the detection of combined genetic alterations (in particular, 5 out of 24 PKD cases were combined with membranopathies), and the diagnosis of PKD in a high percentage of cases suspected of CDA (50%) or in a low percentage of unexplained anemias (4%). NGS also enabled the correct identification of two XH cases, initially suspected of PKD. The pathogenic variants detected in Spain are consistent with those previously reported, with the exception of the lower percentage of R510Q.
Keyword(s): Pyruvate kinase deficiency