Clinical utility of next-generation sequencing in the diagnosis of hereditary haemolytic anaemias.
Hereditary haemolytic anaemias are genetically and phenotypically heterogeneous disorders characterized by increased red cell destruction, with consequences ranging from innocuous to severe life-threatening anaemia. Diagnostic laboratories endeavour to assist clinicians reach the exact patient diagnosis by using tests principally based on morphological and biochemical techniques. However, these routine studies may be inconclusive, particularly in newborn infants and when transfusions have recently been administered. Large numbers and size of the potentially involved genes also impose a practical challenge for molecular diagnosis using routine sequencing approaches. To overcome these diagnostic shortcomings, we have utilized next-generation sequencing to provide a high-throughput, highly sensitive assay. We developed a panel interrogating 28 genes encoding cytoskeletal proteins and enzymes with sequencing coverage of the coding regions, splice site junctions, deep intronic and regulatory regions. We then evaluated 19 samples, including infants with unexplained extreme hyperbilirubinaemia and patients with transfusion-dependent haemolytic anaemia. Where possible, inheritance patterns of pathogenic mutations were determined by sequencing of immediate relatives. We conclude that this next-generation sequencing panel could be a cost-effective approach to molecular diagnosis of hereditary haemolytic anaemia, especially when the family history is uninformative or when routine laboratory testing fails to identify the causative haemolytic process.
Department of Pathology, University of Utah and ARUP Laboratories, Salt Lake City, UT, USA.
Archana M
AM
Agarwal
Associated Regional and University Pathologists (ARUP) Laboratories, Salt Lake City, UT, USA.
Roberto H
RH
Nussenzveig
Associated Regional and University Pathologists (ARUP) Laboratories, Salt Lake City, UT, USA.
Noel S
NS
Reading
Department of Pathology, University of Utah and ARUP Laboratories, Salt Lake City, UT, USA.
Jay L
JL
Patel
Department of Pathology and Laboratory Medicine, Western University, London, ON, Canada.
Nikhil
N
Sangle
Department of Pathology, University of Utah and ARUP Laboratories, Salt Lake City, UT, USA.
Mohamed E
ME
Salama
Division of Hematology, Department of Medicine, University of Utah Health Sciences, Salt Lake City, UT, USA.
Josef T
JT
Prchal
Department of Pathology, University of Utah and ARUP Laboratories, Salt Lake City, UT, USA.
Sherrie L
SL
Perkins
Division of Hematology/Oncology, Department of Pediatrics, University of Utah School of Medicine, Salt Lake City, UT, USA.
Hassan M
HM
Yaish
Divisions of Neonatology and Hematology Oncology, University of Utah School of Medicine, Salt Lake City, UT, USA.
Robert D
RD
Christensen
D000293:Adolescent; D000328:Adult; D000745:Anemia, Hemolytic, Congenital / Q000175:diagnosis* / Q000235:genetics; D002648:Child; D002675:Child, Preschool; D003598:Cytoskeletal Proteins / Q000235:genetics; D004798:Enzymes / Q000235:genetics; D040461:Gene Components / Q000235:genetics; D059014:High-Throughput Nucleotide Sequencing / Q000191:economics / Q000379:methods*; D006801:Humans; D006933:Hyperbilirubinemia, Hereditary / Q000175:diagnosis; D007223:Infant; D007231:Infant, Newborn; D025202:Molecular Diagnostic Techniques / Q000191:economics / Q000379:methods; D009154:Mutation; D055815:Young Adult
D003598:Cytoskeletal Proteins; D004798:Enzymes
RBC membrane defect; haemolytic anaemia; hereditary anaemias; hereditary spherocytosis; next generation sequencing