A Possible Intrinsic Mechanism For Clonal Expansion Of Pnh Abnormal Cells

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired hematopoietic stem cell disorder that causes clonal expansion of Glycosylphosphatidylinositol (GPI)-anchor deficient cells. A somatic mutation of PIG-A that encodes a subunit of the enzyme complex in the GPI-anchor synthesis results in deficiency of GPI-anchored proteins. Two lines of evidence suggest that second abnormality is involved in clonal expansion of GPI-anchor deficient cells. Pig-a disrupted hematopoietic stem cells in mouse and PIG-A deficient cells in normal human individuals did not clonally expand Two possible mechanisms for clonal expansion have been proposed One is an immunoselection mechanism in that PNH cells are more resistant to immunological attack than normal cells because of the deficiency of GPI-anchored proteins. The other is an intrinsic mechanism in that PNH cells have a second mutation responsible for clonal expansion like benign tumors.We reported that all of the GPI-anchor deficient cells in a patient with PNH carried both abnormality of chromosome 12, 46XX, t(12; 12)(q13; q15) and a mutation of PIG-A. We hypothesized that the abnormality of chromosome 12 is causally related to clonal expansion of PNH cells in this patient. In order to map the breakpoints and identify genes located there, we established hybrid cell lines by fusing GPI-anchor deficient monocytes of the patient with HPRT-deficient mouse myeloma cells and selected hybrid cell lines carrying each abnormal chromosome 12 by analyzing microsatellite markers. We determined the breakpoints by testing the presence of STS markers, Southern blotting and the inverse PCR method. We found that a 9.3Mbp fragment derived from 12q13-q15 region in one chromosome 12 was inserted into q15 region in the other chromosome 12. So, the chromosome abnormality was not a translocation but an insertion of a region from q13 to q15 and should be shown 46XX, ins(12; 12)(q15; q13q15).We have identified a gene located in one of the breakpoints. I will discuss its possible involvement in clonal expansion of PNH abnormal cells.