Molecular Complete Remission of Clonal Hematopoiesis in Paroxysmal Nocturnal Hemoglobinuria (PNH) after Syngeneic Transplantation.

PNH is a clonal hematopoietic stem cell disorder and transplantation is potentially curative. However, the pathobiology of PNH is different from malignant clonal myelopathies such as acute leukemia, and the optimal approach to transplant is unresolved. Somatic mutation of PIGA underlies deficiency of glycosyl phosphatidylinositol anchored proteins (GPI-APs) that accounts for the hemolytic anemia of PNH but does not explain the clonal expansion required for clinical manifestation of the disease to become apparent. Recently we identified two patients whose PIGA mutant cells had a concurrent rearrangement of chromosome 12 that resulted in ectopic expression of HMGA2, the architectural transcription factor deregulated in many benign tumors. These observations suggest that aberrant HMGA2 expression, in concert with mutant PIGA, accounted for clonal hematopoiesis in these patients and support the view of PNH as a benign tumor of the bone marrow. One of the patients (a 31 y.o. female) underwent syngeneic SCT because of persistent transfusion-dependent hemolysis and thromboembolic complications that occurred despite prophylactic anticoagulation. At presentation, flow cytometry showed that 95% of PMN were GPI-AP deficient. Marrow analysis revealed erythroid hyperplasia and a karyotype of 46,XX,ins(12)(p13q15q13) in 20/20 (100%) metaphases. The cytogenetic abnormality was shown to be an intrachromosomal insertion that deregulated expression of HMGA2. A 14 bp deletion in exon 2 of PIGA was identified in PMN DNA. Quantitative PCR showed mutant PIGA and der(12) in 91.8% and 93.9%, respectively of bone marrow cells, demonstrating that the two mutations coexisted in the same cells. Neither mutant PIGA nor der(12) was identified in hematopoietic elements of the syngeneic donor, confirming their somatic origin. Donor CD34⁺ cells were mobilized with filgrastim to yield 5 × 10⁶ cells/kg recipient weight. Because of the complex molecular abnormalities and the anticipated absence of graft vs. tumor effect, a preparative regimen of targeted busulfan (1mg/kg × 16 doses) and cyclophosphamide (50 mg/kg × 4 doses) was given. Engraftment occurred on Day 9 with no subsequent hemolysis. At 6 and 15 months post-transplant, high sensitivity flow cytometry showed no GPI-AP deficient RBCs or PMNs. Moreover, PCR revealed no mutant PIGA or der(12) post-transplant, demonstrating, for the first time, a molecular complete remission of PNH following SCT. These results reveal important concepts in PNH pathobiology and transplant efficacy. 1) As the donor was a syngeneic twin, graft vs. tumor effect was not required for eradication of the PNH clone. 2) Prior to transplant, essentially all hematopoiesis was derived from the double mutant clone, but the marrow ablative regimen completely eradicated the clone without the need for pretransplant cytoreduction. Together, these observations add further support to the concept of PNH as a benign tumor of the marrow and suggest a rational approach to SCT.