Expression of HMGA2 In Blood Cells From Patients with Paroxysmal Nocturnal Hemoglobinuria

Abstract 4242

Paroxysmal nocturnal hemoglobinuria (PNH) is caused by a somatic mutation of PIG-A gene in one or few hematopoietic stem cells and subsequent clonal expansion of mutant stem cells. It is known that PIG-A mutation is insufficient to account for the clonal expansion required for clinical manifestation of PNH. We proposed a 3-step model of PNH pathogenesis. Step 1 involves the generation of a GPI-deficient hematopoietic stem cell by somatic mutation of PIG-A gene. Step 2 involves the immunological selection of GPI-deficient hematopoietic stem cells. Based on the close association of PNH with aplastic anemia, it has been suggested that the selection pressure is immune mediated. However, in spite that over 60% of patients with aplastic anemia have subclinical population of GPI-deficient hematopoietic cells at diagnosis, only 10% develop clinical PNH, suggesting that steps-1 and 2 are insufficient to cause PNH. Under immune mediated selection pressure, GPI-deficient cells not only survive, but also must proliferate much more frequently than usual to compensate for anemia. This elevated proliferation rate may increase a chance that additional mutations are acquired, in turn leads to step 3. Step 3 involves a second somatic mutation that bestows on PIG-A mutant stem cell a proliferative phenotype. According to this hypothesis, we searched for the candidate gene for step 3. We reported 2 patients with PNH whose PIG-A mutant cells had an acquired rearrangement of chromosome12, generating the break within the 3′ untranslated region in HMGA2. This gene encodes an architectural transcription factor which is deregulated in many benign mesenchymal tumors (Blood. 2006 vol.108 no.13, p4232). Based on these, we consider HMGA2 as a candidate gene, ectopic expression of which causes proliferation of PIG-A mutant cells. We reported that the expression of HMGA2 in peripheral blood from PNH patients was significantly higher than that from normal volunteers (relative mRNA expression, 4.8±2.4 vs 1.3±0.3, p<0.05) but this was not the case in the bone marrow. We investigated whether over expression of HMGA2 really causes the clonal expansion using the mouse model. We transduced the mouse bone marrow cells with retrovirus vectors, pMYs-HMGA2-IRES-EGFP or pMY-IRES-EGFP as a control, and transplanted them into lethally irradiated mice. The percentage of HMGA2 expressing cells in peripheral blood cells of each lineage from transplanted mice gradually increased during 4-months' observation, suggesting that over expression of HMGA2 caused clonal expansion of multipotent hematopoietic cells. This result is consistent with a recent report on the gene therapy of beta-thalathemia that clonal expansion of rescued hematopoietic cells occurred due to lentivaral insertion into and ectopic expression of HMGA2 (Science. 2009, 326, p1468). Investigation of the somatic mutation which causes upregulation of HMGA2 is being conducted. The dysregulation of Wnt pathway is one of the candidate mechanisms (Blood. 2009, vol.114, no.22, p786). These results are consistent with our 3-step model of PNH pathogenesis, that is, clonal expansion is caused not only by survival advantage from immunological attack but also by benign-tumor-like proliferation.