Immune pressure on hematopoietic stem cells can drastically expand glycosylphosphatidylinositol-deficient clones in paroxysmal nocturnal hemoglobinuria

Abstract

Abstract Background Paroxysmal nocturnal hemoglobinuria (PNH) is a rare hematological disease characterized by intravascular hemolysis, thrombosis, and bone marrow (BM) failure. Although PNH is caused by excessive proliferation of hematopoietic stem cell (HSC) clones with loss of function mutations in phosphatidylinositol N-acetylglucosaminyltransferase subunit A (PIGA) gene, the factor that drives the expansion of PNH clones remains elusive. Case presentation: We present a 26-year-old female who presented with hemolytic anemia, thrombocytopenia, and leukopenia. Flow cytometry analysis of peripheral blood showed that 71.9% and 15.3% of the granulocytes and erythrocytes were glycosylphosphatidylinositol (GPI)(-). The patient was diagnosed with PNH with BM failure. Deep-targeted sequencing covering 369 different genes of sorted GPI(-) granulocytes revealed three different PIGA mutations (p.I69fs, variant allele frequency (VAF) 24.2%; p.T192P, VAF 5.8%; p.V300fs, VAF 5.1%) and no other mutations. She received six cycles of eculizumab and oral cyclosporine. Although the serum lactate dehydrogenase level decreased, the patient remained dependent on red blood cell transfusion. Six months after diagnosis, she received a syngeneic bone marrow transplant (BMT) from a genetically identical healthy twin, following an immune ablative conditioning regimen consisting of cyclophosphamide 200 mg/kg and rabbit anti-thymocyte globulin 10 mg/kg. After 4 years, the patient’s blood count remained normal without any signs of hemolysis. However, the peripheral blood contained 0.2% GPI(-) granulocytes, and the three PIGA mutations that had been detected before BMT persisted at proportions similar to those before transplantation (p.I69fs, VAF 36.1%; p.T192P, VAF 3.7%; p.V300fs, VAF 8.6%). Conclusions The PNH clones whose proportion had increased excessively before BMT decreased but persisted at low percentages for more than 4 years after the immunoablative conditioning regimen followed by syngeneic BMT. These findings indicate that immune pressure on HSCs, which caused BM failure before BMT, was sufficient for PIGA-mutated HSCs to clonally expand to cause PNH.