ALBERT

Description: Rearrangements involving the MLL gene at chromosome 11q23 are associated with leukemia and are present in up to 70% of infant leukemias. Loss of heterozygosity (LOH) has been shown for anonymous polymorphic markers at 11q23 in adult leukemias. To study LOH at theMLL locus, we have identified two new polymorphic microsatellite markers: a GAA repeat (mllGAAn) in intron 6 of theMLL gene and a GA (mllGAn) repeat in the 5′ flanking region of the gene, approximately 2 kb upstream of the translation initiation codon. The heterozygosity index of mllGAAn is 0.54, which renders it useful for analyzing LOH. We screened two groups of leukemia patients to study LOH at the mllGAAn marker. Group A (n = 18) was selected on the basis of presentation before 18 months. Cytogenetic and reverse transcription-polymerase chain reaction analysis showed that 9 of these 18 children had translocations involving MLL. No LOH was observed. Group B (n = 36) were randomly selected children who had presented with leukemia between 1993 and 1994. Cytogenetic analysis of this group showed a variety of different chromosomal abnormalities. LOH was shown in 9 of 20 individuals (45%) who were informative. Microsatellite instability (MSI) was demonstrated in 1 of 18 individuals in group A and 5 of 36 individuals (13.9%) in group B. MSI and LOH were observed simultaneously in three individuals. Loss of an allele was confirmed in one individual by fluorescence in situ hybridization. Individuals with MSI or LOH at mllGAAn were selected for analysis at anonymous polymorphic markers D11S1364 and D11S1356, which flank the MLL gene. No LOH or MSI was observed at these markers in those individuals who were informative. These results show that LOH at the MLL gene locus is a common event during leukemogenesis. Furthermore, the presence of MSI at this locus suggests that the region is a hotspot for genetic instability.

Description: The optimal approach for stem cell transplantation in children with immunodeficiency has yet to be determined. Conditioning therapy is necessary for reliable engraftment and full immune reconstitution; however, the beneficial effect of cytoreductive conditioning is counterbalanced by increased short- and long-term treatment-related toxicity. Whether bone marrow transplantation with a nonmyeloablative preparative regimen was sufficient for the establishment of donor immune reconstitution, with the resultant correction of disease phenotype, was investigated. Eight patients with severe immunodeficiency states underwent T-cell replete bone marrow transplantation from a human leukocyte antigen-matched unrelated (n = 6) or sibling (n = 2) donor with nonmyeloablative conditioning using a fludarabine–melphalan–anti-lymphocyte globulin-based regimen. All patients had severe organ dysfunction that precluded transplantation with conventional conditioning. All patients were engrafted with predominantly donor hematopoiesis, and the duration of neutropenia was brief. Significant acute graft-versus-host disease (GVHD) did not develop, but one patient had limited chronic GVHD. One patient died of disease recurrence, and 3 have stable, mixed chimerism. At a median follow-up of 1 year, all patients have had good recovery of CD3+ T-cell numbers, and 6 of 7 evaluable patients have normal phytohemagglutinin stimulation indices. The rate of immune reconstitution is comparable with that of historical controls undergoing standard myeloablative protocols. Two patients with CD40 ligand deficiency now show significant expression, and a patient with adenosine deaminase deficiency has improved deoxy adenosine triphosphate metabolites. In summary, it has been demonstrated that nonmyeloablative stem cell transplantation permits rapid engraftment from both sibling and unrelated donors with minimal toxicity even in the presence of severe organ dysfunction. If long-term immune reconstitution of patients treated with this protocol is demonstrated, it is believed this approach might offer significant advantages compared with standard protocols by combining adequate immune reconstitution with reduced short- and long-term toxicity.

Description: The optimal approach for stem cell transplantation in children with immunodeficiency has yet to be determined. Conditioning therapy is necessary for reliable engraftment and full immune reconstitution; however, the beneficial effect of cytoreductive conditioning is counterbalanced by increased short- and long-term treatment-related toxicity. Whether bone marrow transplantation with a nonmyeloablative preparative regimen was sufficient for the establishment of donor immune reconstitution, with the resultant correction of disease phenotype, was investigated. Eight patients with severe immunodeficiency states underwent T-cell replete bone marrow transplantation from a human leukocyte antigen-matched unrelated (n = 6) or sibling (n = 2) donor with nonmyeloablative conditioning using a fludarabine–melphalan–anti-lymphocyte globulin-based regimen. All patients had severe organ dysfunction that precluded transplantation with conventional conditioning. All patients were engrafted with predominantly donor hematopoiesis, and the duration of neutropenia was brief. Significant acute graft-versus-host disease (GVHD) did not develop, but one patient had limited chronic GVHD. One patient died of disease recurrence, and 3 have stable, mixed chimerism. At a median follow-up of 1 year, all patients have had good recovery of CD3+ T-cell numbers, and 6 of 7 evaluable patients have normal phytohemagglutinin stimulation indices. The rate of immune reconstitution is comparable with that of historical controls undergoing standard myeloablative protocols. Two patients with CD40 ligand deficiency now show significant expression, and a patient with adenosine deaminase deficiency has improved deoxy adenosine triphosphate metabolites. In summary, it has been demonstrated that nonmyeloablative stem cell transplantation permits rapid engraftment from both sibling and unrelated donors with minimal toxicity even in the presence of severe organ dysfunction. If long-term immune reconstitution of patients treated with this protocol is demonstrated, it is believed this approach might offer significant advantages compared with standard protocols by combining adequate immune reconstitution with reduced short- and long-term toxicity.